Automated assay processing unit

Abstract

The present disclosure provides automated assay processing units comprising a plurality of components for processing a sample and analyzing the components of the sample when the sample is present in a consumable. The present disclose also provides automated assay processing units for processing large numbers of consumables simultaneously.

Description

AUTOMATED ASSAY PROCESSING UNITCROSS-REFERENCE TO RELATE APPLICATIONS

[0001] This application claims priority benefit to the filing date of U.S. Patent Application Serial No. 63 / 718,500, filed on November 8, 2024, the disclosure of which application is herein incorporated by reference in its entirety.INTRODUCTION

[0002] Diagnostic methods typically involve obtaining and analyzing biological samples from subjects and processing the biological samples for detection and / or quantification of diagnostic analytes, such as biomarkers indicative of diseases. In clinical laboratories, various patient samples are needed to be analyzed for different analytes. Such analyses is conventionally performed using separate analysis systems for different analytes or types of analytes. This increases cost, equipment footprint, required space, energy, reagents, and consumables.SUMMARY

[0003] Certain aspects of the disclosure provide assay processing units that can analyze different analytes or types of analytes thereby reducing cost, equipment, space, energy, reagents, and consumables.

[0004] In certain aspects, the disclosure provides assay processing units that comprise multiple modules, each module configured for engaging a consumable. Thus, in some cases, the analysis units can engage with multiple consumables. Such analyses between multiple consumables can be performed simultaneously.

[0005] In some cases, assay processing units are a multi-functional or configured to allow analysis of different analytes or different types of analytes. For example, assay processing units can perform one or more of a plurality of assays, such as nucleic acid (NA) amplification and detection, immunoassay, etc.

[0006] Further aspects of the disclosure also provides methods of analyzing one or more samples in the automated assay processing units disclosed herein.BRIEF DESCRIPTION OF THE FIGURES

[0007] The disclosure is best understood from the following detailed description when read in conjunction with the accompanying drawings. It is emphasized that, according to common practice, the various features of the drawings are not to-scale. On the contrary, the dimensions ofthe various features are arbitrarily expanded or reduced for clarity. Included in the drawings are the following figures.

[0008] FIG. 1 depicts an exemplary flow diagram of an automated analysis that might be performed on an embodiment of automated assay processing unit according to the present disclosure.

[0009] FIGS. 2A-2B depict exemplary flow diagrams of automated analyses that might be performed on an embodiment of automated assay processing system according to the present disclosure.

[0010] FIGS. 3A-3Z depict aspects of an exemplary embodiment of an automated assay processing system according to the present disclosure. “TPH” as used in FIG. 3W refers to “Tests per hour”.

[0011] FIG. 4A-4B depicts an experiment in which a first population of green particles and a second population of red particles were used to detect beta-human chorionic gonadotropin (B- hCG).

[0012] FIG. 5 depicts exemplary results for the detection of Alpha-fetoprotein (AFP) in low analyte samples.

[0013] FIG. 6 depicts exemplary results for the detection of multiplex AFP and Total Prostate - Specific Antigen (T-PSA).

[0014] FIG. 7 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0015] FIG. 8 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0016] FIG. 9 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0017] FIG. 10 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0018] FIG. 11 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0019] FIG. 12 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0020] FIG. 13 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0021] FIG. 14 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0022] FIG. 15 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0023] FIG. 16 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0024] FIG. 17 depicts exemplary results for the detection of multiplex AFP and T-PSA.

[0025] FIG. 18 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats. “SMP” as disclosed in FIG. 18 refers to “sample”.

[0026] FIG. 19 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats. “SMP” as disclosed in FIG. 18 refers to “sample”.

[0027] FIG. 20 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0028] FIG. 21 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0029] FIG. 22 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0030] FIG. 23 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0031] FIG. 24 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0032] FIG. 25 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0033] FIG. 26 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0034] FIG. 27 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0035] FIG. 28 depicts exemplary results for B-hCG microparticles with varying levels of Ab coats.

[0036] FIG. 29 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.

[0037] FIG. 30 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.

[0038] FIG. 31 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.

[0039] FIG. 32 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.

[0040] FIG. 33 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.

[0041] FIG. 34 depicts exemplary results for a Traumatic Brain Injury (TBI) assay.DETAILED DESCRIPTION

[0042] Aspects of the present disclosure provide automated assay processing units and methods of using the automated assay processing units.

[0043] In certain embodiments, the automated assay processing units of the present disclosure comprise a first module comprising a sample processing component comprising a consumable holder; a second module comprising an aspiration and dispense component; a third module comprising a mixing component; and a fourth module comprising an optics component, wherein the sample processing component is configured to translate the consumable holder to a plurality of positions within the sample processing component, the first, second, third and fourth modules are operably connected and configured to interface with a consumable located on the consumable holder.

[0044] In certain embodiments, the automated medium-capacity assay processing units of the present disclosure comprise two or more fluid addition stations, two or more wash stations, two or more imaging stations, and a consumable mover.

[0045] In certain embodiments, the automated high-capacity assay processing units of the present disclosure comprise four or more fluid addition stations, two or more wash stations, wherein each wash station comprises a first carousel encircled by a second carousel, four or more imaging stations, and a consumable mover.

[0046] Various hardware components and associated features are described throughout this disclosure. Automated assay processing units may include any of the features and respective hardware components described or combinations thereof. However, some features and components may make more practical sense on one particular aspect over the other, depending on particular design considerations.

[0047] Before the present systems and methods are described in greater detail, it is to be understood that the present disclosure is not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.

[0048] The terms “automated assay processing units,” “units,” or “systems,” or the like terms may be used interchangeably throughout the disclosure, and each refers to automated assay processing units, standard automated assay processing units, automated medium-capacity assay processing units or automated high-capacity assay processing units, for example, as depicted in FIG. 3S.

[0049] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the devices and methods. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the devices and methods, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the devices and methods.

[0050] Certain ranges are presented herein with numerical values being preceded by the term “about.” The term “about” is used herein to provide literal support for the exact number that it precedes, as well as a number that is near to or approximately the number that the term precedes. In determining whether a number is near to or approximately a specifically recited number, the near or approximating un-recited number may be a number which, in the context in which it is presented, provides the substantial equivalent of the specifically recited number.

[0051] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present devices and methods, representative devices and methods are now described.

[0052] The present disclosure may be understood more readily by reference to the following detailed description of desired embodiments and the examples included therein. In the following specification and the claims which follow, reference will be made to a number of terms which shall be defined to have the following meanings.

[0053] Although specific terms are used in the following description for the sake of clarity, these terms are intended to refer only to the particular structure of the embodiments selected for illustration in the drawings, and are not intended to define or limit the scope of the disclosure. In the drawings and the following description below, it is to be understood that like numeric designations refer to components of like function.

[0054] The term “comprising” is used herein as requiring the presence of the named component and allowing the presence of other components. The term “comprising” should be construed to include the term “consisting essentially of’ and “consisting of.” The “consisting essentially of’ allows the presence of the named component(s), along with other component which do not change the function / structure of the named component(s). The “consisting of’ allows the presence of the named component(s), along with any adhesives or other bonding means for attaching the listed component(s).

[0055] Numerical values should be understood to include numerical values which are the same when reduced to the same number of significant figures and numerical values which differ from the stated value by less than the experimental error of conventional measurement technique of the type described in the present application to determine the value.

[0056] All ranges disclosed herein are inclusive of the recited endpoint and independently combinable (for example, the range of “from 2 grams to 10 grams” is inclusive of the endpoints, 2 grams and 10 grams, and all the intermediate values). The endpoints of the ranges and any values disclosed herein are not limited to the precise range or value; they are sufficiently imprecise to include values approximating these ranges and / or values.

[0057] The modifier “about” used in connection with a quantity is inclusive of the stated value and has the meaning dictated by the context. When used in the context of a range, the modifier “about” should also be considered as disclosing the range defined by the absolute values of the two endpoints. For example, the range of from about “2 to about 10” also discloses the range “from 2 to 10.” The term “about” may refer to plus or minus 10% of the indicated number. Forexample, “about 10%” may indicate a range of 9% to 11%, and “about 1” may mean from 0.9- 1.1.

[0058] It should be noted that many of the terms used herein are relative terms. For example, the terms “upper” and “lower” are relative to each other in location, i.e., an upper component is located at a higher elevation than a lower component in a given orientation, but these terms can change if the component is flipped. The terms “inlet” and “outlet” are relative to a fluid flowing through them with respect to a given structure, e.g., a fluid flows through the inlet into the structure and flows through the outlet out of the structure.

[0059] The terms “horizontal” and “vertical” are used to indicate direction relative to an absolute reference, i.e., ground level. However, these terms should not be construed to require structures to be absolutely parallel or absolutely perpendicular to each other. For example, a first vertical structure and a second vertical structure are not necessarily parallel to each other. The terms “top” and “bottom” are used to refer to surfaces where the top is always higher than the bottom relative to an absolute reference, i.e., the surface of the earth. The terms “upwards” and “downwards” are also relative to an absolute reference; upwards is always against the gravity of the earth while downwards is always towards the gravity of the earth.

[0060] The term “parallel” should be construed in its lay sense of two surfaces that maintain a generally constant distance between them, and not in the strict mathematical sense that such surfaces will never intersect when extended to infinity.

[0061] “Microbead” and “microparticle” are used herein interchangeably and refer to a substantially spherical solid support. The microbead or microparticle is a substantially spherical solid support that is influenced by a magnetic field such that the magnetic field can attract or repulse the microparticle or magnetic particle. A microbead or microparticle may occupy or settle in an array of wells, such as, for example, in an array of wells in a detection module. The microparticle and microbead may contain at least one specific binding member that binds to an analyte of interest and at least one detectable label. Alternatively, the microparticle and microbead may contain a first specific binding member that binds to the analyte and a second specific binding member that also binds to the analyte and contains at least one detectable label.

[0062] "Non-fnnctional bead," "helper bead," and "assisting particle" are used interchangeably and refers to a substantially spherical assisting solid support, that is larger in diameter than a microparticle, which is configured to be chemically inert with respect to other components of an assay. As used herein, an assisting particle refers to a spherical particle which generally does not chemically interact with other particles (including a microparticle, conjugate, and / or reagent), but which is magnetic or paramagnetic. In certain exemplary embodiments, assisting particle may be coated so as to chemically interact with interferents, that is, any materials which would interferewith assay or analysis of an analyte of interest within the targeted sample. In such embodiments, the assisting particles can also improve binding efficiency of the microparticles including, for the purpose of illustration and not limitation, by binding with interferents. Additionally and alternatively, the shape of a solid support can be roughly spherical, though not limited to such shapes.

[0063] The assisting solid supports can be larger in diameter than the other support mediums within the storage region and configured so as to not chemically interact with any other components within the mixing region. Specifically, the diameter of the assisting solid supports (e.g. helper beads) can be at least 1 %, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least about 11 %, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 50%, at least 75%, at least 100%, at least 150%, at least 200%, at least 250%, at least 300%, at least 350%, at least 400%, at least 450%, at least 500%, at least 550%, at least 600%, at least 650%, at least 700%, at least 750%, at least 800%, at least 850%, at least 900%, at least 950%, at least 1000% greater or larger than the diameter of other support mediums (e.g., microparticles).

[0064] Isolating an analyte refers to removing away the analyte from additional molecules present with the analyte, for example, other components in a biological sample comprising the analyte.

[0065] All publications and patents cited in this specification are herein incorporated by reference as if each individual publication or patent were specifically and individually indicated to be incorporated by reference and are incorporated herein by reference to disclose and describe the methods and / or materials in connection with which the publications are cited. The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present disclosure is not entitled to antedate such publication by virtue of prior disclosure. Further, the dates of publication provided may be different from the actual publication dates, which may need to be independently confirmed.

[0066] It is noted that, as used herein and in the appended claims, the singular forms “a”, “an”, and “the” include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as “solely,” “only” and the like in connection with the recitation of claim elements or use of a “negative” limitation.

[0067] As will be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present devices and methods. Anyrecited method can be carried out in the order of events recited or in any other order which is logically possible.AUTOM TED ASSAY PROCESSING UNITS

[0068] As summarized above, aspects of the present disclosure provide automated assay processing units and methods of using the automated assay processing units. Automated assay processing units may also be referred to as automated assay processing systems.

[0069] As discussed above, diagnostic methods involve analyzing biological samples for detection and / or quantification of diagnostic analytes, such as biomarkers indicative of diseases. In a laboratory environment, diagnostic methods include analyzing various patient samples for different analytes or biomarkers.

[0070] Conventionally, analysis systems are designed for analyzing a specific analyte or a type of analyte. For example, systems for analyzing nucleic acids are designed for amplification and detection of nucleic acids and, therefore, such analysis systems can only be used for analyzing nucleic acids.

[0071] Similarly, systems for analyzing proteins or peptides, such as immunoassay devices, are specifically designed for detecting proteins or peptides and, in many cases, even specific proteins or peptides. Therefore, such systems can only be used for analyzing proteins or peptides or even specific proteins or peptides.

[0072] Thus, diagnostic analyses are conventionally performed using separate analysis systems for different analytes or types of analytes. Therefore, a typical diagnostic laboratory houses many analysis systems, each designed to specifically analyze one or only a few analytes or types of analytes. This increases operating cost, for example, cost associated with equipment, space, energy, reagents, and consumables.

[0073] The automated assay processing unit described herein allows analyzing multiple analytes using a single / universal system, multiple systems and, optionally, a single / universal consumable, such as a cartridge, thereby reducing costs associated with equipment, space, energy, reagents, and consumables.

[0074] Moreover, because of the modularity and multiplex capability of the automated assay processing unit, the unit provides ease of use. Conventionally, specific training is required for performing unique assays. However, minimal operator training would be necessary for operating the automated assay processing unit disclosed herein as a single consumable or a handful of consumables are capable of conducting multiple types of analyses.

[0075] The automated assay processing unit of the present disclosure is capable of being joined with other automated assay processing units to increase the number of assays performed at onceand the total throughput of the number of assays performed in an hour. Because each automated assay processing unit is able to perform a specific assay independently of other automated assay processing units, the assays may be configured to have: different durations of specific assay steps, different numbers of steps performed in a given assay, the ability to have each automated assay processing unit to perform different assays, etc. The combination of multiple automated assay processing units also allows for an ease of maintenance because any given assay processing unit may be deactivated and repaired or maintained without the need to shutdown, turnoff, or alter the activity the other automated processing units that are not undergoing maintenance.

[0076] FIG. 1 depicts an exemplary immunoassay flow diagram that may be performed on an embodiment of an automated assay processing unit according to the present disclosure. Flow diagram 100 depicts the steps involved in conducting an exemplary immunoassay. Such steps are broadly categorized as a first immunocomplex reaction 110 (i.e., Immunocomplex Reaction (IR)- Part 1 or IR1), followed by immunocomplex reaction 120 (i.e., Immunocomplex Reaction (IR)- Part 2 or IR2), and finally followed by enzymatic reaction 130 (i.e., Enzymatic Reaction (ER)). As seen in the figure, enzymatic reaction 130 comprises capture image stack steps following the seeding of microparticles into microwells and the sealing of the microwells, including Capture Image Stack 1 step 140. Each capture image stack comprises a plurality of constituent steps. Image Stack Capture Activities 150 shows an exploded view of Capture Image Stack 1 step 140 with a plurality of constituent steps related to adjusting LED configurations and evaluating focus of the imaging conducted at the step.

[0077] FIGS. 2A-2B depict exemplary flow diagrams of the timing, e.g., staging, and order of exemplary immunoassays performed on an embodiment of an automated assay processing system according to the present disclosure. Flow diagram 200 depicts the steps involved in conducting an exemplary analysis in conjunction with a corresponding “stage” or “position” of the automated assay processing system; i.e., a “stage” or “position” of a sample processing component of a first module of an automated assay processing system, as described herein. Flow diagram 200 shows analysis steps performed at Stage 1 at step 210; i.e., the analysis steps performed sample processing component of a first module of an automated assay processing system positions a consumable present in a consumable holder in a first position; i.e., translates the consumable holder into a first position. Generally, steps 210 of flow diagram 200 performed at Stage 1 correspond to loading materials, samples, reagents and the like onto one or more consumables present in the automated assay processing system.

[0078] After performing the analysis and / or pre-analytical steps set forth in step 210, the automated assay processing system transitions a consumable present in the consumable holder from a first position (i.e., Stage 1) to a second position corresponding to Stage 2, at which steps220 are performed, by moving the consumable through Transition 1 at step 215. When a step is listed in a transition, e.g., transition 1 215 or transition 2 225, the step may be performed before the transition or after the transition. Generally, steps 220 of flow diagram 200 performed at Stage 2 correspond to performing immunocomplex reactions, such as Immunocomplex Reaction - Part 1 (IR1) and / or Immunocomplex Reaction - Part 2 (IR2).

[0079] After performing the analysis steps set forth in step 220, the automated assay processing system transitions a consumable present in the consumable holder from a second position (i.e., Stage 2) to a third position corresponding to Stage 3, at which steps 230 are performed, by moving the consumable through Transition 2 at step 225. Generally, steps 230 of flow diagram 200 performed at Stage 3 correspond to performing enzymatic reaction (ER), including, for example, image acquisition and enzymatic reaction incubation.

[0080] FIG. 2B depicts flow diagram 250, which is an alternative analysis flow diagram, which begins with steps 260 at Stage 1 before transitioning in Transition 1 265 to steps 270 at Stage 2 before transitioning in Transition 2 275 to steps 280 at Stage 3. When a step is listed in a transition, e.g., transition 1 265 or transition 2 275, the step may be performed before the transition or after the transition.

[0081] FIGS. 3A-3Z depict aspects of an exemplary embodiment of an automated assay processing system according to the present disclosure. Automated assay processing system 300 comprises a first module with a sample processing component 310. Automated assay processing unit 300 may be referred to as a standard automated assay processing unit. Sample processing component 310 comprises a consumable holder 320 for holding a consumable (not shown), as such are described herein. In an embodiment, the sample processing component 310 moves the consumable into one or more positions, e.g., at least two, at least three positions. Automated assay processing system 300 further comprises a second module. In embodiments, the second module comprises an aspiration and dispense (ADS) component 330. Automated assay processing system 300 further comprises a third module. The third module comprises a mixing component 340. Automated assay processing system 300 further comprises a fourth module. The fourth module comprises optics component 350. First, second, third and fourth modules are operably connected together such that each module is available to interface with consumable 325 positioned on consumable holder 320. Sample processing component 310 is configured to translate consumable holder 320 to a plurality of different positions or locations within sample processing component 310. In embodiments, sample processing component 310 is configured to translate consumable holder 320 to a plurality of positions, wherein each position or location is configured such that aspects of the first module, the second module, the third module and the fourth module caninterface with or interact with or operate on consumable 325 and contents thereof, e.g., sample, reagents, buffers or the like.

[0082] Embodiments of automated assay processing unit further comprise a degassing component. Any convenient degassing unit may be employed. In some embodiments, the degassing component comprises a vacuum component to remove air from one or more reagents present in the consumable. In some embodiments, the degassing component comprises a cooling component to cool one or more reagents present in the consumable thereby reducing the air present in one or more reagents in the consumable.First magnet:

[0083] Additionally or alternatively, in embodiments, sample processing component 310 of the first module comprises first magnet 321. In embodiments, first magnet 321 is a retractable magnet such that magnet can be retracted into magnet pocket 321a. Additionally or alternatively, first magnet 321 may be rotated relative to consumable holder 320 or configured such that first magnet 321 can be rotated, e.g., dynamically rotated. In some embodiments, first magnet 321 is not rotated dynamically and is only rotated by un-securing, e.g., detaching, the first magnet from the sample processing component and re-securing, e.g., re-attaching, the first magnet in a different orientation. In embodiments, sample processing component 310 is configured such that first magnet 321 is positioned beneath or below consumable holder 320. When magnet 321 is a retractable magnet, it may be configured such that it is positioned proximate to consumable holder 320 in an un-retracted state and distal to consumable holder 320 in a retracted state. By “proximate," it is meant that first magnet 321 is positioned sufficiently near or close to consumable holder 320 or positioned such that a distance between first magnet 321 and consumable holder320 is such that first magnet exerts a magnetic field on consumable 325 present in consumable holder 320 such that one or more particles present in consumable 325 are affected by the magnetic field of first magnet 321. By “distal," it is meant that first magnet 321 is positioned sufficiently far or away from consumable holder 320 or positioned such that a distance between first magnet321 and consumable holder 320 is such that the first magnet does not exert a meaningful magnetic field on consumable 325 present in consumable holder 320 such that one or more particles present in consumable 325 are not affected by the magnetic field of first magnet 321. In some cases, automated assay processing system 300 is configured such that first retractable magnet 321 is encased or enclosed or housed in a sleeve or pocket 321a in the retracted state wherein sleeve or pocket 321a, e.g., a laminated structure on the sleeve or pocket, shields first retractable magnet 321 thereby reducing the local magnetic field of first magnet 321. That is, sleeve or pocket 321a is configured to shield the magnetic field of first magnet 321 when first magnet 321 is retractedinto sleeve or pocket 321a such that the magnetic field of first magnet 321 is sufficiently minimized viz-a-viz particles or substances present in consumable 325 held in consumable holder 320. In some embodiment, sleeve or pocket 321a is made of a material that shields the consumable from the magnetic field. The material may be any material that influences magnetic fields including, without limitation, iron, nickel, cobalt, steel, brass, copper, aluminum, alloys containing ferromagnetic metals, etc.Functional positions of sample processing component:

[0084] Additionally or alternatively, in embodiments, automated assay processing system 300 is configured such that sample processing component 310 is configured to translate consumable holder 320 to at least three positions, comprising first position 326, second position 327 and third position 328, within sample processing component 320. The translation of the consumable holder may be in a single dimension, e.g., a x-dimension or a y-dimension, or may be in multiple dimensions, e.g., a x-dimension and a y-dimension.

[0085] In embodiments, first position 326 is a loading / unloading position. That is, first position 326 is located so as to enable loading or unloading of, e.g., sample, reagents, buffers or the like, onto consumable 325. For example, in some cases, first position 326 may be positioned such that sample, reagents, buffers or the like may be loaded into or unloaded out of one or more wells, receptacles or the like present on consumable 325. For example, first position 326 may be positioned such that one or more wells, partitions, receptacles or the like, present on consumable 325 are exposed or uncovered, or are aligned with loading or unloading mechanisms, such that loading / unloading onto consumable 325 can occur. For example, when consumable holder 325 is in first position 326, aspects of the second module or the third module may be activated or controlled or otherwise engaged with consumable 325 in order to performs steps related to loading and / or unloading contents, e.g., sample, reagent, buffers or the like, onto or out of consumable 325. Additionally or alternatively, first position 326 may be used as a loading / unloading position for the unloaded or unloading of the consumable such that when the consumable holder is present in first position 326, the consumable may be loading into or unloading out of the consumable holder. In some embodiments, the sample processing component 310 comprises a first heating element in the first position beneath the consumable holder that increases the temperature of, maintains the temperature of, or reduces the temperature of the consumable when the consumable is present in the first position. The first heating element is described in further detail below.

[0086] Tn embodiments, second position 327 is associated with immunocomplex reaction - part 1(IR1) and immunocomplex reaction - part 2 (IR2) as well as mixing and dispensing, e.g., sample, reagents, buffers or the like, present on consumable 325. In embodiments, IRl-related processinginvolves performing one or more steps depicted in block 110 of FIG. 1. In embodiments, Unrelated processing involves performing one or more steps depicted in block 120 of FIG. 1. In embodiments, sample processing component 320 translates consumable holder 325 into second position 327 that is a location such that, for example, aspects of the second module and / or the third module can interface with the contents of consumable 325. For example, when consumable holder 325 is in second position 327, aspects of the second module may be activated or controlled or otherwise engaged with consumable 325 in order to performs steps related to aspirating and / or dispensing materials, e.g., sample, reagent, buffers or the like, onto or out of consumable 325. Additionally or alternatively, when consumable holder 325 is in second position 327, the third module may be activated or controlled or otherwise engaged with consumable 325 in order to perform steps related to mixing materials, e.g., sample, reagents, buffers or the like, present on consumable 325. Additionally or alternatively, when consumable holder 325 is in second position 327, one or more modules of system 300, e.g., aspects of the second module or the third module, may be activated or controlled or otherwise engaged with consumable 325 in order to perform steps related to performing IR1 and / or IR2 with respect to the contents, e.g., sample, reagents, buffers or the like, of consumable 325. In some embodiments, the sample processing component 310 comprises a second heating element in the second position beneath the consumable holder that increases the temperature of, maintains the temperature of, or reduces the temperature of the consumable when the consumable is present in the first position. The second heating element is described in further detail below.

[0087] In embodiments, third position 328 is associated with imaging aspects of consumable 325 or the contents thereof. In embodiments, third position 328 is also associated with performing steps related to enzymatic reaction (ER) or a portion thereof with respect to the contents, e.g., sample, reagents, buffers or the like, present on consumable 325. In embodiments, ER -processing involves performing one or more steps depicted in block 130 of FIG. 1. In some embodiments, only ER-processing steps following oil sealing are performed in the third position. In some embodiments, seeding, sealing, and waste disposal are performed in the third position. In embodiments, sample processing component 320 translates consumable holder 325 into third position 328 that is a location such that, for example, the fourth module can interface with consumable 325. For example, when consumable holder 325 is in third position 328, aspects of system 300, such as the second module, the third module and / or the fourth module, may be activated or controlled or otherwise engaged with consumable 325 in order to perform steps related to performing or conducting one or more enzymatic reactions (ER) with respect to the contents of consumable 325. Additionally or alternatively, when consumable holder 325 is in third position 328, the fourth module may be activated or controlled or otherwise engaged withconsumable 325 in order to image aspects of consumable 325, including the contents thereof. In some instances, when consumable holder 325 is in third position 328, the fourth module may be engaged with consumable 325 through an imaging clamping mechanism described below. In some cases, third position 327 corresponds to a final position with respect to one or more reactions or assays having been completed when consumable 325 is positioned in third position 327 such that imaging of consumable 325 occurs in connection with evaluating the results of such reactions or assays.Compression elements:

[0088] In embodiments, sample processing component 310 comprises one or more compression elements 329. Such compression elements 329 may be configured to engage consumable holder 320 or to engage consumable 325 when consumable 325 is present in consumable holder 320 and the consumable 325 and / or consumable holder 325 are in the second position. In embodiments, compression elements 329 are fingers. In some cases, fingers 329 are metallic strips configured to act as springs. In embodiments, fingers 329 are configured to engage consumable holder 320. In embodiments, fingers 329 are configured to engage consumable 325 present in consumable holder 320. In embodiments, fingers 329 are configured to apply a downward force on consumable holder 320. In embodiments, fingers 329 are configured to apply a downward force on consumable 325 and consumable holder 320. In embodiments, fingers 329 are configured to clamp consumable 325 between fingers 329 and consumable holder 320. In embodiments compression elements 329 are configured to engage consumable holder 320 or consumable 325 when consumable holder 320 or consumable 325 is in second position 327. In embodiments, compression elements 329 are configured to hold consumable 325 in a relatively fixed position when consumable holder 325 is in second position 327 allowing or enabling or facilitating one or more aspects of system 300 can operate on consumable 325 in order to perform one or more aspects of IR1, IR2 or mixing and dispensing-related steps. In some embodiments compression elements 329 still have contact with part of the consumable 325 in holder 320 when the holder 320 has placed consumable 325 into position 328. The downward force provided by the compression on the consumable provides the benefit of ensuring uniform engagement of the consumable with the consumable holder and the second heating element positioned underneath the consumable holder in the second position thereby increasing the efficiency and stability of heating of the consumable and the reagent, sample, etc., deposited therein.Heating elements:

[0089] In embodiments, sample processing component 310 comprises a first heating element 345 located beneath consumable holder 320 in the first position. In embodiments, the first heating element is configured to heat consumable 325 present in consumable holder 320. In some cases, the first heating element is configured to heat consumable 325 to a specified temperature, such as to a first temperature. The first temperature may be any convenient temperature, such as a temperature selected to facilitate one or more steps of loading, unloading, 1R1, IR2, mixing and dispensing, ER or imaging-related steps. In some cases, the first temperature is a temperature that is 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, 100 C or greater. In some embodiments, the first temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C, about 55-60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than 100 C. In some cases, sample processing component 310 is configured such that the first heating element heats consumable 325 when consumable holder 320 is positioned in first position 326. In embodiments, the first heating element is configured to heat the entire consumable 325 to a first temperature or any convenient part or subset or aspect of consumable 325, such as one or more wells or receptacles of consumable 325, to a first temperature. In embodiments, any convenient heating element or apparatus or device, such as any convenient commercially available heating element, such as, for example, resistive heating elements, may be utilized for the second heating element.

[0090] In embodiments, sample processing component 310 comprises a second heating element 346 located beneath consumable holder 320 in the second position. In embodiments, the second heating element is configured to heat consumable 325 present in consumable holder 320. In some cases, the second heating element is configured to heat consumable 325 to a specified temperature, such as to a second temperature. The second temperature may be any convenient temperature, such as a temperature selected to facilitate one or more steps of loading, unloading, IR1, IR2, mixing and dispensing, ER or imaging-related steps. In some cases, the second temperature is a temperature that is about 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the second temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C,about 55-60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than about 100 C. In some cases, sample processing component 310 is configured such that the second heating element heats consumable 325 when consumable holder 320 is positioned in second position 327. In embodiments, the second heating element is configured to heat the entire consumable 325 to a second temperature or any convenient part or subset or aspect of consumable 325, such as one or more wells or receptacles of consumable 325, to a second temperature. In embodiments, any convenient heating element or apparatus or device, such as any convenient commercially available heating element, such as, for example, resistive heating elements, may be utilized for the second heating element.

[0091] In embodiments, the first temperature may differ from the second temperature, i.e., such that consumable 325, or aspects thereof, is heated to a first temperature when in first position 326 and heated to a different temperature in second position 327. First temperature may be greater than second temperature by any convenient amount, such as by about 0 C, about 5 C, about 10 C, about 15 C, about 20 C or greater. First temperature may be less than second temperature by any convenient amount such as by 0 C, about 5 C, about 10 C, about 15 C, about 20 C or greater. In other embodiments, the first temperature may be the same as the second temperature, i.e., such that consumable 325, or aspects thereof, is heated to a first temperature when in first position 326 and heated to the same temperature in second position 327.

[0092] In some embodiments, the first heating element and the second heating element are separated in that there is no cross-talk between the first heating element and the second heating element. By “separate” it is meant that the first heating and the second heating element are not connected. The first heating element and the second heating element may be separated like the first position 326 and the second position 327 are separated in FIG. 3J. Separate heating elements provide the benefit of customizable temperatures per position.

[0093] In embodiments, sample processing component 310 comprises a first temperature sensor. In embodiments, sample processing component 320 may be configured such that the first temperature sensor senses a temperature, e.g., of consumable 325, or aspects of consumable, or consumable holder 320, or aspects of consumable holder 320, when consumable holder 320 is located in the first position 326 or the ambient temperature in the first position. In embodiments, sample processing component 320 comprises a second temperature sensor. In embodiments, sample processing component 320 may be configured such that the second temperature sensor senses a temperature, e.g., of consumable 325, or aspects of consumable, or consumable holder 320, or aspects of consumable holder 320, when consumable holder 320 is located in second position 327 or the ambient temperature in the second position. In embodiments, sample processing component 320 comprises a third temperature sensor. In embodiments, sampleprocessing component 320 may be configured such that the third temperature sensor senses a temperature, e.g., of consumable 325, or aspects of consumable, or consumable holder 320, or aspects of consumable holder 320, when consumable holder 320 is located in third position 328 or the ambient temperature in the third position. In embodiments, any convenient temperature sensor, such as any convenient commercially available temperature sensor, such as, for example, one or more ambient temperature sensors, may be utilized for the first and second temperature sensors.Imaging clamping mechanism:

[0094] In embodiments, sample processing component 310 comprises a raising element configured to engage consumable holder 320 or to engage consumable 325 present in consumable holder. In some embodiments, the raising element is the imaging component 352 that is capable of being raised to the same height or above the sample processing component 310. The raising element may be further configured to engage consumable holder 325 or consumable by raising consumable holder 320 or consumable 325. In some cases, the raising element may be configured to engage consumable holder 320 or consumable with aspects of fourth module comprising aspects of an optics component 350. In some cases, the raising element is configured to engage consumable holder 320 or consumable 325 when consumable holder 320 is in third position 328. In embodiments, the raising element may be configured to engage consumable 325 and apply an upward force. For example, the raising element may be configured to engage consumable 325 and push consumable 325 upwards such that consumable is clamped between the raising mechanism and a clamping component 401a and 401b. The clamping component 401a and 401b provides a downward force in opposition to the upward force of the raising element such that the combination forces holds the consumable in place. The raising element in combination with the clamping element provides the benefit of stabilizing the consumable during the imaging process. In embodiments, the raising element may be configured to hold consumable 325 in a fixed position such that aspects of fourth module 350, such as one or more optics components 350 can be engaged or controlled to collect one or more images of consumable 325 or the contents thereof. In embodiments, the raising element may be configured to hold consumable 325 substantially in a fixed position for a period of time during which fourth module 350 images one or more aspects of consumable 325, such as one or more sections of consumable 325 over time. In embodiments, the raising element may comprise one or more constituent components, such as, for example, one or more springs or one or more clamps or one or more motors or one or more cams. In embodiments, the raising element raises consumable holder 320 or consumable 325 whenconsumable holder 320 or consumable 325 physically contacts aspects of the fourth module, such as optics component 350 thereof.

[0095] In some cases, in conjunction with the raising element, consumable holder 320 or consumable 325 physically contacts an alignment datum 353 present on optics component 350 of the fourth module. In embodiments, alignment datum 353 is a peg or a post protruding from a horizontal surface of optics component 350 of the fourth module with a fixed position in space relative to imaging components, such as, for example, a camera, of optics component 350. In embodiments, alignment datum 353 is configured to engage or abut or act as a stop for consumable holder 320 or consumable 325 present thereon, such that when consumable holder 320 or consumable 325 engages alignment datum 353, the contents, e.g., wells, of consumable 325 are in a fixed position relative to, e.g., a camera, of optics component 350.Aspiration and dispense component;

[0096] In embodiments, the second module comprises an aspiration and dispense (ADS) component 330. In embodiments, aspiration and dispense component 330 is configured to engage consumable 325 present on consumable holder 320 such that aspects of aspiration and dispense component 330 can aspirate materials, e.g., sample, buffers, reagents or the like, from consumable 325 and / or dispense materials, e.g., sample, buffers, reagents or the like, to consumable 325. In embodiments, the aspiration and dispense component comprises one or more pumps, such as aspiration pump 331a and oil pump 331b. Aspiration pump 331a may be fluidically connected to an aspiration nozzle 403. In embodiments, aspiration pump 331a is configured to aspirate a hydrophilic liquid through the aspiration nozzle 403 when consumable holder 320 is in second position 327, i.e., when consumable holder 320 locates consumable 325 in a second position 327 such that aspects of the aspiration and dispense component can access consumable 325, i.e., one or more wells, well arrays, or partitions of consumable 325.

[0097] In embodiments, the aspiration and dispense component comprises a hydrophilic liquid nozzle fluidically connected to a hydrophilic liquid pump. In some cases, the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle, wherein the consumable holder is in the second position.

[0098] Oil pump 331b may be configured to dispense hydrophobic liquid, e.g., oil, and may be connected to a nozzle, e.g., fluidically connected to a nozzle configured to dispense fluid into one or more aspects of consumable 325, e.g., one or more wells, well arrays, or partitions of consumable 325. Tn embodiments, the aspiration and dispense component comprises a hydrophobic liquid nozzle 402 fluidically connected to a hydrophobic liquid pump 331b. In somecases, the hydrophobic liquid pump 33 lb is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle 402 when the consumable holder is in the second position.

[0099] In embodiments, the aspiration and dispense component is positioned above sample processing component 320. In embodiments, third module 330 comprising the aspiration and dispense component is positioned above sample processing component 320. That is, third module 330 comprising the aspiration and dispense component is positioned to access wells or partitions of consumable 325 held in consumable holder 320 of sample processing component 320. In some embodiments, the third module is capable of being moved such that the aspiration nozzle 403 and the hydrophobic liquid nozzle 402 is in proximity to a desired region of the consumable such that the aspiration nozzle can remove a desired liquid from the consumable and the hydrophobic liquid nozzle can deposit the hydrophobic liquid into a desired location of the consumable. In some embodiments, the consumable and consumable holder are capable of being moved such that the aspiration nozzle 403 and the hydrophobic liquid nozzle 402 is in proximity to a desired region of the consumable such that the aspiration nozzle can remove a desired liquid from the consumable and the hydrophobic liquid nozzle can deposit the hydrophobic liquid into a desired location of the consumable.

[0100] In some cases, the aspiration and dispense component comprises aspiration pump 331a and hydrophobic liquid pump 331b, and aspiration pump 331a is configured to aspirate hydrophilic liquid and hydrophobic liquid pump 33 lb is configured to dispense the hydrophobic liquid. In some cases, aspiration pump 331a and hydrophobic liquid pump 331b are configured to aspirate and dispense, respectively, simultaneously. In some cases, aspiration pump 331a and hydrophobic liquid pump 331b are configured to aspirate and dispense, respectively, when consumable holder 320 is in the second position 327.Second magnet:

[0101] In embodiments, the aspiration and dispense component comprises second magnet 332. In embodiments, the mixing component comprises second magnet 332. In embodiments, second magnet 332 is a retractable magnet such that magnet can be retracted into a magnet pocket. Additionally or alternatively, second magnet 332 may be rotated relative to consumable holder 320 or configured such that second magnet 332 can be rotated, e.g., dynamically rotated. In embodiments, aspiration and dispense component 330 is configured such that second magnet 332 is positioned above consumable holder 320, i.e., second magnet 332 is an “overhead” magnet. When magnet 332 is a retractable magnet, it may be configured such that it is positioned proximate to consumable holder 320 in an un-retracted state and distal to consumable holder 320 in a retracted state. By “proximate,” it is meant that second magnet 332 is positioned sufficiently nearor close to consumable holder 320 or positioned such that a distance between second magnet 332 and consumable holder 320 is such that second magnet 332 exerts a magnetic field on consumable 325 present in consumable holder 320 such that one or more particles present in consumable 325 are affected by the magnetic field of second magnet 332. By “distal,” it is meant that second magnet 332 is positioned sufficiently far or away from consumable holder 320 or positioned such that a distance between second magnet 332 and consumable holder 320 is such that second magnet 332 does not exert a meaningful magnetic field on consumable 325 present in consumable holder320 such that one or more particles present in consumable 325 are not affected by the magnetic field of second magnet 332. In some cases, automated assay processing system 300 is configured such that second retractable magnet 332 is encased or enclosed or housed in a sleeve or pocket in the retracted state wherein such sleeve or pocket shields second retractable magnet 332 thereby reducing the local magnetic field of second magnet 332. That is, such sleeve or pocket is configured to shield the magnetic field of second magnet 332 when second magnet 332 is retracted into such sleeve or pocket such that the magnetic field of second magnet 332 is sufficiently minimized viz-a-viz particles or substances present in consumable 325 held in consumable holder 320. In some embodiment, sleeve or pocket is made of a material that shields the consumable from the magnetic field. The material may be any material that influences magnetic fields including, without limitation, iron, nickel, cobalt, steel, brass, copper, aluminum, alloys containing ferromagnetic metals, etc. In embodiments, second magnet 332 is positioned above consumable holder 320 or consumable 325 present therein when consumable holder 320 is in second position 327. That is, when consumable holder is in second position 327, second magnet 332 is aligned with consumable holder 320 and consumable 325 present therein.

[0102] In some embodiments in which automated sample processing system 300 comprises each of first magnet 321 and second magnet 332, such first and second magnets are horizontally offset from each other. That is, first magnet 321 and second magnet 332 are horizontally offset from each other within system 300. That is, first magnet 321 and second magnet 332 are sufficiently separated from each other to prevent magnetic fields from such magnets from interacting with each other or to minimize interaction between magnetic fields of each such magnet. In some cases, first magnet 321 and second magnet 332 are configured such that first magnet 321 and second magnet 332 cannot both be in an un-retracted state at the same time. That is, first magnet321 and second magnet 332 are interlocked to prevent both first magnet 321 and second magnet 332 from being in an un-retracted state at the same time, similarly, to prevent magnetic fields from such magnets from interacting with each other or to minimize interaction between magnetic fields of each such magnet. The first magnet and the second magnet may be un-retracted and retracted in an alternating fashion such that alternating magnet fields may be applied to the consumable.The consumable and consumable holder may be moved between the first magnet and the second magnet in an alternating fashion, e.g., from the first magnet to the second magnet in a horizontal position, in order to alternate the magnetic fields on the consumable. In some cases, first magnet 321 and second magnet 332 are configured such that first magnet 321 and second magnet 332 can both be in an un-retracted state at the same time such that the first magnet and the second magnet can exert magnetic fields on the consumable simultaneously.Reagent pumps and nozzles:

[0103] In embodiments, aspiration and dispense component 330 comprises one or more reagent pumps fluidically connected to one or more reagent nozzles. In embodiments, system 300 may comprise reagent pumps in addition to, i.e., separate from, aspiration pump 331a and / or oil pump 331b. In embodiments, the one or more reagent pumps may comprise, or may be fluidically connected to, reagent nozzle 333. In some cases, the one or more reagent pumps may comprise, or may be fluidically connected to, one or more reagent nozzles 333.

[0104] In embodiments, the one or more reagent pumps are configured to dispense fluid through the one or more reagent nozzles. For example, the one or more reagent pumps may be configured to dispense reagent through the one or more reagent nozzles. Any convenient fluid, including any convenient reagent, capable of being dispensed through reagent pump may be dispensed, such as a wash buffer. By dispense, it is meant dispense into consumable 325 present on consumable holder 320. For example, it is meant, dispense fluid, such as a reagent, into a well or partition of consumable 325. Further, it is meant to dispense a specified amount of fluid. For example, about 1-10, about 10-20, about 20-30, about 30-40, about 40-50, about 50-60, about 60-70, about 70-80, about 80-90, about 90-100, about 100-110, about 110-120, about 120-130, about 130-140, about 140-150, about 150-160, about 160-170, about 170-180, about 180-190, about 190-200, about or greater than about 200 pL may be dispensed. In some embodiments, the reagent pumps and reagent nozzles dispense reagents at high precision. For example, the reagent pumps and reagent nozzles may dispense reagents within about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 1 1 , about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, or about 20 pL of the desired dispense volume. In embodiments, the one or more reagent pumps are configured to dispense fluid through the one or more reagent nozzles when consumable holder 320 is in second position 327. That is, when consumable holder 320 is positioned in second position 327, such one or more reagent nozzles are aligned with one or more wells, well arrays, or partitions of consumable 325, such that such reagent nozzles can dispense fluid, e.g., reagent, into a well or partition of consumable 325. In other words, system 300 is configured such that when consumable 325 is position in second position 327 of consumableholder 320, such one or more reagent nozzles of aspiration and dispense component 330 align with consumable 325, in particular an aspect of consumable 325 configured to receive fluid, e.g., reagent.

[0105] In embodiments, mixing component 340 comprises one or more reagent pumps fluidically connected to one or more reagent nozzles. In embodiments, such reagent pumps may comprise one or more buffer pump 341. In embodiments, system 300 may comprise reagent pumps in addition to, i.e., separate from, buffer pump 341. In embodiments, the one or more reagent pumps may comprise, or may be fluidically connected to, wash buffer dispense 333. In some cases, the one or more reagent pumps may comprise, or may be fluidically connected to, one or more reagent nozzles 333.

[0106] In embodiments, the one or more reagent pumps are configured to dispense fluid through the one or more reagent nozzles. For example, the one or more reagent pumps may be configured to dispense reagent through the one or more reagent nozzles. Any convenient fluid, including any convenient reagent, capable of being dispensed through reagent pump may be dispensed. In embodiments, the one or more reagent pumps may be configured to dispense a reagent through wash buffer dispense 333. By dispense, it is meant dispense into consumable 325 present on consumable holder 320. For example, it is meant, dispense fluid, such as a reagent, into a well or partition of consumable 325. Further, it is meant dispense a specified amount of fluid. In embodiments, the one or more reagent pumps are configured to dispense fluid through the one or more reagent nozzles when consumable holder 320 is in second position 327. That is, when consumable holder 320 is positioned in second position 327, such one or more reagent nozzles are aligned with one or more wells, well arrays, or partitions of consumable 325, such that such reagent nozzles can dispense fluid, e.g., reagent, into a well or partition of consumable 325. In other words, system 300 is configured such that when consumable 325 is position in second position 327 of consumable holder 320, such one or more reagent nozzles of aspiration and dispense component 330 align with consumable 325, in particular an aspect of consumable 325 configured to receive fluid, e.g., reagent.Mixing motor and arm;

[0107] In embodiments, mixing component 340 comprises mixing motor 342 and engagement arm 343 that is configured to engage with consumable 325 present in consumable holder 320. The mixing motor may be a range of different motors, including, without limitation, a stepper motor, a brushless DC motor, a solenoid, etc. In some embodiments, the mixing motor is a voice coil. In some cases, engagement arm 343 engages with a portion of consumable 325. For example, engagement arm 343 may engage with a dedicated position of consumable 325, such as a slot ordepression or cutout or extended ann 343. In some cases, engagement arm 343 engages with a dedicated portion of consumable 325 in order to mix or vibrate or agitate or otherwise disrupt one or more aspects of the portion of consumable 325. For example, engagement arm 343 may engage with a dedicated portion of consumable 325 in order to mix only a subset of the wells or partitions of consumable 325. In other words, engagement arm 343 may engage with a dedicated portion of consumable 325 in order to isolate mixing to only a subset of consumable 325. In an embodiment, the end 344 of engagement arm 343 is shown engaging with a portion of consumable 325 in FIG. 3P. In some embodiments, the engagement arm engages with a portion of the consumable that is not a portion dedicated to mixing and provides localized vibration to the consumable.

[0108] In other cases, engagement arm 343 engages with entire consumable 325. In such cases, engagement arm 343 may be configured to mix or vibrate or agitate or otherwise disrupt the entire consumable 325 or the entire contents of consumable 325, i.e., the each well or partition of consumable 325.

[0001] In embodiments, the engagement arm is attached to a chamber in the shape of a cube, rectangle, or cylinder comprising a diaphragm on the interior and an opening at one end with a suction cup or a gasket. The suction cup or the gasket contacts a portion of the consumable. The diaphragm may be moved or oscillated by a deflection amount that compresses the air in the chamber. The compression in the air results in movement in the fluid consumable thereby mixing the fluid. The chamber comprises one or more openings that allow pressure relief in the chamber when the diaphragm is moving or oscillating. The chamber may be according to the pressure mixing device disclosed in US Patent Application No. 18 / 763,854.

[0002] The diaphragm of the sample mixing device may be made of a range of different materials including, without limitation, piezoelectric material, metal, a compliant material that deforms under heat or pressure, etc. The diaphragm of the sample mixing device may be moved or oscillated in a number of different ways. In some embodiments, the diaphragm is made of a piezoelectric material. In some embodiments, the piezoelectric material is moved or oscillated by applying an electric current to the piezoelectric material. In some embodiments, the diaphragm is metal. In some embodiments, the metal is moved or oscillated by turning a magnetic field on and off. In some embodiments, the diaphragm is made of a compliant material that deforms under heat or pressure. In some embodiments, the compliant material is moved by apply air pressure to the compliant material. In some embodiments, the compliant material is moved by applying heat to the compliant material. In some embodiments, the compliant material is moved by applying acoustic waves to the compliant material. In some embodiments, the compliant material is moved by applying a mechanical force to the compliant material.

[0109] In embodiments, mixing component 340 comprises a ballistic mixing device. In some cases, the ballistic mixing device is a pipettor. That is, mixing component 340 may be configured to activate or control a pipettor to mix or vibrate or agitate or otherwise disrupt one or more aspects, e.g., one or more wells, well arrays, or partitions of consumable 325.Imaging component:

[0110] In embodiments, optics component 350 comprises imaging component 352. In embodiments, optics component 350 comprises a secondary illumination component 393 in addition to the imaging component 352. The secondary illumination component 393 can be positioned above or below the consumable. The secondary illumination component 393 may be used to obtain a forward scatter image of the consumable or a back scatter image of the consumable. In some embodiments, the secondary illumination component 393 is positioned above the consumable. In some embodiments, the secondary illumination component is positioned below the consumable. In some embodiments, the secondary illumination component comprises a second printed circuit board comprising one or more LEDs. In embodiments, imaging component 352 is positioned beneath consumable holder 320. In embodiments, imaging component 352 is positioned beneath consumable holder 320 when consumable holder 320 is in the third position 328. That is, when consumable holder 320 is positioned in third position 328, imaging component 352 is aligned with consumable 325, such as aligned with one or more wells, well arrays, or partitions of consumable 325, such that imaging component 352 can image consumable 325 or image specified portions thereof, such as one or more wells, well arrays, or partitions of consumable 325. In embodiments, imaging component 352 is positioned above consumable holder 320 when consumable holder 320 is in the third position 328. That is, when consumable holder 320 is positioned in third position 328, imaging component 352 is aligned with consumable 325, such as aligned with one or more wells, well arrays, or partitions of consumable 325, such that imaging component 352 can image consumable 325 or image specified portions thereof, such as one or more wells, well arrays, or partitions of consumable 325. In other words, system 300 is configured such that when consumable 325 is position in third position 328 of consumable holder 320, such imaging component 352 of optics component 350 align with consumable 325, in particular an aspect of consumable 325, such as one or more wells, well arrays, or partitions, the contents of which are to be imaged. The alignment of the consumable and the imaging component may be achieve by engaging the consumable with an alignment datum.[0011 1 ] Tn certain embodiments, imaging component 352 comprises an illumination component. Illumination components of the instant disclosure include any excitation light source sufficient to generate excitation light that, after passing through the optics block, excites a fluorophore presentin the sample which, in turn, produces emission light. Accordingly, the subject illumination components of the instant disclosure will vary and may include but are not limited to e.g., lamps, lasers, light emitting diodes (LED), and the like. The illumination component may be configured to facilitate imaging consumable 325 by illuminating consumable 325 or one or more portions of consumable 325. In some cases, the illumination component comprises one or more lighting elements. For example, the illumination component may comprise one or more LEDs 355. In some cases, the illumination component is enclosed or encased or present within enclosed illumination chamber 354. In some cases, enclosed illumination chamber 354 comprises opening or window or aperture 356. In embodiments, opening or window or aperture 356 is positioned beneath consumable holder 320 when consumable holder is in third position 328. That is, when consumable holder 320 is positioned in third position 328, such opening or window or aperture 356 is aligned with consumable 325 or one or more wells, well arrays, or partitions of consumable 325, such that opening or window or aperture 356 allows consumable 325 or aspects thereof to be illuminated and / or imaged by illumination component, e.g., LEDs 355, and / or imaging component. In other words, system 300 is configured such that when consumable 325 is positioned in third position 328 of consumable holder 320, such aperture 356 of optics component 350 aligns with consumable 325, or an aspect of consumable 325, such as a well or partition thereof, such that consumable can be illuminated and / or imaged through aperture 356.

[0112] In some embodiments, the imaging component comprises an objective lens 425 positioned below the PCB 358. In some embodiments, the imaging component comprises a second aperture 420 positioned below the objective lens 425. In some embodiments, the imaging component comprises an emission filter 421 positioned below the second aperture 420. The emission filter may be a dual band emission filter. In some embodiments, the imaging component comprises an imaging lens 423 positioned below the emission filter 421. In some embodiments, the imaging component comprises a field flattening lens 424 positioned below the imaging lens 423. The field flatten lens may be a lens that expands the usable field of view. In some embodiments, the imaging component comprises a sensor 426 positioned below the field flattening lens 424.

[0113] The optics component comprises a secondary illumination component and an imaging component 352 comprising an illumination component. The secondary illumination component 393 allows for the identification of objects, e.g., microparticles in wells, in a consumable in the absence of fluorescence. The objects are identified through forward or back scatter imaging produced from the secondary illumination component that may be position above or below the consumable. The secondary illumination component is aligned with the emission wavelengths (per emission wavelength) for the purposes of obtaining either a back or front scatter (respectively) image.

[0114] In some instances, the illumination component and / or secondary illumination component contains one or more LEDs including but not limited to e.g., two or more LEDs, three or more LEDs, four or more LEDs, one LED, two LEDs, three LEDs, four LEDs, five LEDs, six LEDs, seven LEDs, eight LEDs, nine LEDs, ten LEDs, etc. In some instances, an illumination component and / or secondary illumination component containing four LEDs may contain two pairs of identical LEDs or one pair of LEDs of a first wavelength and a second pair of LEDs of a second wavelength. In instances where a plurality of LEDs is employed, any useful arrangement of the LEDs may find use in the illumination component and / or secondary illumination component including but not limited to e.g., linear arrangement, staggered arrangement, arrayed (e.g., “checker-board”) arrangement, a circular arrangement, and the like. Useful LEDs of the subject disclosure will vary, e.g., based on the particular assay to be performed by the device the optical, electrical or physical constraints of the device and the like. The illumination component and / or secondary illumination component of the present disclosure is capable of changing the intensity of each LED or the channel type of each.

[0115] LEDs useful in an illumination component and / or secondary illumination component of the subject imaging component may include but are not limited to e.g., LEDs with a peak minimum wavelength (X) in nanometers (nm) of between 350 and 750 nm, including but not limited to e.g., between 350 and 450, between 350 and 400, between 400 and 450, between 450 and 550, between 450 and 500, between 500 and 550, between 550 and 650, between 550 and 600, between 600 and 650, between 650 and 750, between 650 and 700, between 700 and 750, about 400 nm, about 580 nm, about 470 nm, about 628 nm, about 528 nm, about 674 nm, and the like.

[0116] LEDs useful in an illumination component and / or secondary illumination component of the subject imaging component may include but are not limited to e.g., LED emitters with a peak maximum wavelength (X.) in nanometers (nm) of between 350 and 750 nm, including but not limited to e.g., between 350 and 450, between 350 and 400, between 400 and 450, between 450 and 550, between 450 and 500, between 500 and 550, between 550 and 650, between 550 and 600, between 600 and 650, between 650 and 750, between 650 and 700, between 700 and 750, about 400 nm, about 405 nm, about 592 nm, about 480 nm, about 648 nm, about 542 nm, about 689 nm, and the like.

[0117] In some instances, an illumination component of an imaging component and / or secondary illumination component as described herein may include two or more LEDs. In imaging components having two or more LEDs each LED may be defined as belonging to a channel including but not limited to e.g., a first channel, a second channel, etc. In certain instances, the LEDs of a multi-channel, multi-block system may be configured such that the average differencebetween the peak min / max wavelengths of each pair of LEDs that share an optics block is maximized. For example, in such instances, six LEDs having six different peak min / max wavelengths of A, B, C, X, Y, Z (where A < B < C < X < Y < Z) may be paired in optics blocks as, e.g., A and X, B and Y, C and Z. In such instances, where the average difference between the peak min / max wavelengths of each pair of LEDs that share an optics block is maximized, overlap (i.e., crosstalk) between the emission wavelength of the LEDs in each block may be minimized.

[0118] In some instances, an imaging component may contain two LED emitters of different wavelengths where the distance between the different wavelengths will vary and may range from 5 nm to 300 nm or more including but not limited to e.g., at least about 5 nm apart, at least about 10 nm apart, at least about 15 nm apart, at least about 20 nm apart, at least about 25 nm apart, at least about 30 nm apart, at least about 35 nm apart, at least about 40 nm apart, at least about 45 nm apart, at least about 50 nm apart, at least about 55 nm apart, at least about 60 nm apart, at least about 65 nm apart, at least about 70 nm apart, at least about 75 nm apart, at least about 80 nm apart, at least about 85 nm apart, at least about 90 nm apart, at least about 95 nm apart, at least about 100 nm apart, at least about 105 nm apart, at least about 110 nm apart, at least about 115 nm apart, at least about 120 nm apart, at least about 125 nm apart, at least about 130 nm apart, at least about 135 nm apart, at least about 140 nm apart, at least about 145 nm apart, at least about 150 nm apart, at least about 155 nm apart, at least about 160 nm apart, at least about 165 nm apart, at least about 170 nm apart, at least about 175 nm apart, at least about 180 nm apart, at least about 185 nm apart, at least about 190 nm apart, at least about 195 nm apart, at least about 200 nm apart, not more than about 300 nm apart, not more than about 290 nm apart, not more than about 280 nm apart, not more than about 270 nm apart, not more than about 260 nm apart, not more than about 250 nm apart, not more than about 240 nm apart, not more than about 230 nm apart, not more than about 220 nm apart, not more than about 210 nm apart, not more than about 200 nm apart, not more than about 190 nm apart, not more than about 180 nm apart, not more than about 170 nm apart, not more than about 160 nm apart, not more than about 150 nm apart, not more than about 140 nm apart, not more than about 130 nm apart, not more than about 120 nm apart, not more than about 110 nm apart, not more than about 100 nm apart,, etc.

[0119] In some instances, LEDs of the subject disclosure may be capable of being toggled (i.e., capable being turned on and off, including turned on / off repeatedly). In some instances, the wiring circuitry of an optics block having two or more LEDs is configured or the programing controlling such an imaging component is configured such that only one LED emitter may be toggled on at a time. In such instances, when a first LED of an imaging is toggled on the second LED emitter of the imaging is toggled off and vice versa.

[0120] In some instances, the toggling of LEDs of an imaging component includes a time period where neither LED of the imaging component is toggled on. In some instances, such a time period where neither LED of the imaging component is toggled on is between toggling the toggling off of a first LED and the toggling on of a second LED.

[0121] LEDs of the disclosed the systems, devices, and methods may include physical or electrical components and / or configurations allowing for use or one or more of the noise reduction methods described herein, including but not limited to e.g., components and / or configurations for time division multiplexing, components and / or configurations for frequency division multiplexing, components and / or configurations for spatial separation, and the like.

[0122] Imaging components of the instant disclosure will include a variety of optical elements including but not limited to e.g., lenses, optical filters, mirrors (including e.g., dichroic mirrors), apertures, etc.. For example, in one embodiment, an imaging component of the instant disclosure includes an imaging aperture allowing excitation light (generated by the LED unit to pass to the consumable and emission light to pass back to the imaging component, a reference channel aperture allowing light to pass from the imaging component to the signal processing unit in the reference channel, a measurement channel aperture allowing light to pass from the imaging component to the signal processing unit in the measurement channel.

[0123] Emission light, e.g., from a consumable or a control surface such as a dark target, proceeds through the imaging aperture and the imaging aperture lens. Emission light passing through the detection filter is redirected by a second mirror, referred to herein as the detection channel mirror, up towards the detection (i.e., measurement) channel aperture. After passing through the detection channel aperture lens, emission light proceeds through the detection channel aperture, e.g., into a measurement detector as part of or coupled to an optical signal processor. Light in the reference channel proceeds up through the reference channel aperture lens and out the reference channel aperture, e.g., into a reference detector as part of or coupled to an optical signal processor. In some embodiments, the imaging component additionally includes optical elements for increasing the field of view, e.g., field flattening lenses, increasing the depth of focus and resolution, e.g., apertures, etc.

[0124] The arrangement of the optical elements within the imaging component is not limited to those arrangements specifically depicted and may vary provided the necessary elements for producing the described light paths are included and sufficient to generate, pass and filter excitation and emission light as described herein.

[0125] Excitation filters useful in an imaging component of the instant disclosure include but are not limited to e.g., illuminations filters having a center wavelength (CWL) in nanometers (nm) between 350 and 750 nm, including but not limited to e.g., between about 350 and 450, betweenabout 350 and 400, between about 400 and 450, between about 450 and 550, between about 450 and 500, between about 500 and 550, between about 550 and 650, between about 550 and 600, between about 600 and 650, between about 650 and 750, between about 650 and 700, between about 700 and 750, about 409 nm, about 583 nm, about 475 nm, about 638 nm, about 535 nm, about 690 nm, and the like. Excitation filters useful in an optics block of the instant disclosure also include but are not limited to e.g., illuminations filters having a full width have maximum (FWHM) in nm ranging from 5 nm to 100 nm, including but not limited to e.g., about 5 nm to 10 nm, about 10 nm to 15 nm, about 15 nm to 20 nm, about 20 nm to 25 nm, about 25 nm to 30 nm, about 30 nm to 35 nm, about 35 nm to 40 nm, about 40 nm to 45 nm, about 45 nm to 50 nm, about 50 nm to 55 nm, about 55 nm to 60 nm, about 60 nm to 65 nm, about 65 nm to 70 nm, about 70 nm to 75 nm, about 75 nm to 80 nm, about 80 nm to 85 nm, about 85 nm to 90 nm, about 90 nm to 95 nm, about 95 nm to 100 nm, about 10 nm to 90 nm, about 10 nm to 80 nm, about 10 nm to 70 nm, about 10 nm to 60 nm, about 10 nm to 50 nm, about 10 nm to 40 nm, about 10 nm to 30 nm, about 10 nm to 20 nm, about 20 nm to 90 nm, about 30 nm to 90 nm, about 40 nm to 90 nm, about 50 nm to 90 nm, about 60 nm to 90 nm, about 70 nm to 90 nm, about 80 nm to 90 nm, about 65 nm, about 22 nm, about 36 nm, about 24 nm, about 18 nm, about 25 nm, and the like.

[0126] In some instances an excitation filter useful in an imaging component as described herein may be characterized in having a particular combination of center wavelength (CWL) and full width have maximum (FWHM), including e.g., combinations of the CWL and the FWHM described above. For example, in some instances an excitation filter of the subject disclosure may be characterized as having a 409 nm CWL and a 65 nm FWHM, 583 nm CWL and a 22 nm FWHM, 475 nm CWL and a 36 nm FWHM, 638 nm CWL and a 24 nm FWHM, 535 nm CWL and a 18 nm FWHM, 690 nm CWL and a 25 nm FWHM, and the like.

[0127] Emission filters useful in an imaging component of the instant disclosure include but are not limited to e.g., emission filters having a center wavelength (CWL) in nanometers (nm) between 350 and 750 nm, including but not limited to e.g., between 350 and 450, between 350 and 400, between 400 and 450, between 450 and 550, between 450 and 500, between 500 and 550, between 550 and 650, between 550 and 600, between 600 and 650, between 650 and 750, between 650 and 700, between 700 and 750, about 490 nm, about 617 nm, about 524 nm, about 673 nm, about 565 nm, about 715 and the like. Emission filters useful in an imaging component of the instant disclosure also include but are not limited to e.g., emission filters having a full width have maximum (FWHM) in nm ranging from 5 nm to 100 nm, including but not limited to e.g., about 5 nm to 10 nm, about 10 nm to 15 nm, about 15 nm to 20 nm, about 20 nm to 25 nm, about 25 nm to 30 nm, about 30 nm to 35 nm, about 35 nm to 40 nm, about 40 nm to 45 nm, about 45 nm to 50 nm, about 50 nm to 55 nm, about 55 nm to 60 nm, about 60 nm to 65 nm, about 65 nmto 70 nm, about 70 nm to 75 nm, about 75 nm to 80 run. about 80 nm to 85 nm, about 85 nm to 90 nm, about 90 nm to 95 nm, about 95 nm to 100 nm, about 10 nm to 90 nm, about 10 nm to 80 nm, about 10 nm to 70 nm, about 10 nm to 60 nm, about 10 nm to 50 nm, about 10 nm to 40 nm, about 10 nm to 30 nm, about 10 nm to 20 nm, about 20 nm to 90 nm, about 30 nm to 90 nm, about 40 nm to 90 nm, about 50 nm to 90 nm, about 60 nm to 90 nm, about 70 nm to 90 nm, about 80 nm to 90 nm, 42 nm, 22 nm, 24 nm, 20 nm, 18 nm, 36 nm, and the like.

[0128] In some instances an emission filter useful in an imaging components as described herein may be characterized in having a particular combination of CWL and FWHM, including e.g., combinations of the CWL and the FWHM described above. For example, in some instances an illumination filter of the subject disclosure may be characterized as having a 490 nm CWL and a 42 nm FWHM, 617 nm CWL and a 22 nm FWHM, 524 nm CWL and a 24 nm FWHM, 673 nm CWL and a 20 nm FWHM, 565 nm CWL and a 18 nm FWHM, 715 nm CWL and a 36 nm FWHM, and the like.

[0129] In some cases, one or more LEDs 355 are in proximity to one or more thermistors 357. In some embodiments, LEDs 355 are driven by electronics capable of measuring the instantaneous voltage and current consumed by one, or multiple, of the LEDs for any given LED type / channel, allowing the power consumption of the respective LEDs to be computed. That is, at least one thermistor 357 may be a specified distance away from each LED 355. For example, each LED 355 may be no more than 5 cm or 4 cm or 4 cm or 1 cm or 9 mm or 8 mm or 7 mm or 6 mm or 5 mm or 4 mm or 3 mm or 2 mm or 1 mm or less away from a corresponding thermistor 357.

[0130] Embodiments may comprise any convenient number of LEDs 355, such as, for example, about one, about two, about three, about four, about five, about six, about seven, about eight, about nine, about ten, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 20, about 50 or about 100 or more LEDs. Embodiments may comprise any convenient number of thermistors 357, such as, for example, about one, about two, about three, about four, about five, about six, about seven, about eight, about nine, about ten, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 20, about 50 or about 100 or more.

[0131] In embodiments, LEDs 355 may be arranged in any convenient arrangement. For example,LEDs 355 may be arranged in a substantially symmetric arrangement or a substantially asymmetric arrangement. For example, LEDs 355 may be arranged in a specified pattern with respect to aperture 356. For example, LEDs 355 may be arranged in a circular pattern, such as a circular pattern around aperture 356. In embodiments, thermistors 357 may be arranged in a pattern with respect to LEDs 355. For example, thermistors 357 may be arranged in a circular pattern around aperture 356, where a thermistor 357 is located between every two LEDs 355.Positioning of thermistors between LEDs provides the benefit of being able to monitor the temperature of each LED.Printed circuit board:

[0132] In embodiments, one or more LEDs 355 are present on a printer circuit board (PCB) 358. In some cases, PCB 358 is configured to heat and / or cool to a specified temperature. In embodiments, PCB 358 comprises a plurality of heating and / or elements configured to heat and / or cool to a specified temperature. In embodiments, such heating / cooling elements are resistive elements 359. In some cases, PCB 358 is configured to heat and / or cool each of LEDs 355, enclosed illumination chamber 354, and consumable 325 to a specified temperature. In embodiments, resistive elements 359 of PCB 358 are configured to heat and / or cool each of LEDs 355, enclosed illumination chamber 354, and consumable holder 320 to a specified temperature. In some cases, the specified temperature is a temperature that is 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the heating / cooling elements are configured to maintain the LEDs at a specific temperature. In some cases, the specified temperature is a temperature that is 100 C or lower, such as about 90 C, about 85 C, about 80 C, about 75 C, about 60 C, about 55 C, about 50 C, about 45 C, about 40 C or less. In some embodiments, the heating / cooling elements are configured to maintain the LEDs between 65 and 45 C. In some cases, PCB 358 is configured such that resistive elements 359 heat and / or cool one or more of LEDs 355, enclosed illumination chamber 354, and consumable holder 320 when consumable holder 320 is positioned in third position 328. In embodiments, when consumable holder 320 is positioned in third position 328, resistive elements 359 are aligned with consumable 325 or one or more wells, well arrays, or partitions of consumable 325, such that heat from resistive elements 359 heats and / or cools consumable 325 or aspects thereof to a specified temperature. In other words, system 300 is configured such that when consumable 325 is positioned in third position 328 of consumable holder 320, resistive elements 359 of optics component 350 align with consumable 325, or an aspect of consumable 325, such as a well or partition thereof, such that consumable 325 can be heated / cooled to a specified temperature by resistive elements 359. The heating / cooling elements are able to maintain a specific temperature by maintaining a specific temperature such that when the temperature is above a desired temperature the LEDs are cooled by a lower temperature and when the temperature is below a desired temperature the LEDs are heated by a higher temperature.

[0133] In embodiments, PCB 358 comprises a core. In embodiments, the core may comprise a material or a combination of materials selected to improve heat dissipation from PCB 358 or from resistive elements 359 present on PCB 358. In embodiments, the entire PCB may comprise a material or combination of materials selected to improve heat dissipation from PCB 358 or from resistive elements 359 present on PCB 358. Materials that improve heat dissipation include, without limitation, silver, copper, gold, aluminum, iron, nickel, brass, tungsten, zinc, alloys and combinations thereof, etc. In embodiments, PCB 358 comprises an aluminum core.

[0134] In some embodiments, the illumination component 393 comprises a second PCB comprising one or more LEDs. The PCB may be according to any of the PCBs above. The LEDs may be according to any of the LEDs above.Clamping component;

[0135] In embodiments, optics component 350 comprises clamping component 401a and 401b. In embodiments, optics component 350 comprises clamping component 401a and 401b configured to engage consumable holder 320 or to engage consumable 325 present in consumable holder 320. The clamping component 401a and 401b may be further configured to engage consumable holder 325 or consumable 325 by applying a downward clamping force to consumable holder 320 or consumable 325. In some cases, clamping component 401a and 401b may be configured to engage consumable holder 320 or consumable with aspects of fourth module comprising aspects of optics component 350. In some cases, clamping component 401a and 401b is configured to engage consumable holder 320 or consumable 325 when consumable holder 320 is in third position 328. In embodiments, clamping component 401a and 401b may be a clamping mechanism configured to engage consumable 325 and apply a downward force. For example, clamping component 401a and 401b may be configured to engage consumable 325 and push consumable 325 downwards such that consumable is clamped between clamping component 401a and 401b and one or more aspects of fourth module, such as one or more aspects of optics component 350. In embodiments, clamping component 401a and 401b may be configured to hold consumable 325 in a fixed position such that aspects of fourth module 350, such as one or more aspects of optics components 350, can be engaged or controlled to collect one or more images of consumable 325 or the contents thereof. In embodiments, clamping component 351 may be configured to hold consumable 325 substantially in a fixed position for a period of time during which the fourth module, or optics component 350 thereof, images one or more aspects of consumable 325, such as one or more sections of consumable 325 over time. In embodiments, clamping component 401a and 401b may comprise one or more constituent components, such as, for example, one or more springs or one or more clamps or one or more motors or one or more cams. In embodiments, clamping component401a and 401b depresses (i.e., pushes down on) consumable holder 320 or consumable 325 when consumable holder 320 or consumable 325 physically contacts aspects of the fourth module, such as optics component 350 thereof. In some cases, system 300 is configured such that clamping component 401a and 401b clamp consumable 325 when consumable holder 320 is positioned in third position 328. In embodiments, when consumable holder 320 is positioned in third position 328, clamping component 401a and 401b is aligned with consumable 325 such that clamping component 401a and 401b can apply a downward force on consumable 325. In other words, system 300 is configured such that when consumable 325 is positioned in third position 328 of consumable holder 320, clamping component 401a and 401b of optics component 350 aligns with consumable 325, or an aspect of consumable 325, such that consumable 325 can held in a fixed position, e.g., a reference position abutting datum pin 351 by clamping component 401a and 401b.Consumable;

[0136] Embodiments may further comprise consumable 325. In some cases, consumable 325 is a microfluidic device. In embodiments, consumable 325 comprises a plurality of wells or partitions. In some cases, consumable 325 may comprise about 100 or more, about 200 or more, about 500 or more, about 1000 or more, about 2000 or more, about 3000 or more, about 4000 or more 5000 or more, about 6000 or more, about 7000 or more, about 8000 or more, about 9000 or more, about 10000 or more, about 20000 or more, about 30000 or more, about 40000 or more 50000 or more, about 60000 or more, about 70000 or more, about 80000 or more, about 90000 or more, about 100000 or more, about 200000 or more, about 300000 or more, about 400000 or more, about 500000 or more, about 600000 or more, about 700000 or more, about 800000 or more, about 900000 or more, about or 1000000 or more wells or microwells. In some embodiments, the sample detection zone comprises 100000 or more wells or microwells. The wells or microwells may be dimensioned such that they are capable of holding or more particles. In embodiments, consumable 325 may comprise any convenient material, such as a biocompatible material, such as a relatively inert material or a relatively non-reactive material, such as a plastic, such as a material with a treated surface, such as a non-porous material or treatment. Embodiments of assay processing units are configured to accept and operate on consumables that are identical to consumables utilized in embodiments of medium-capacity assay processing units or automated high-capacity assay processing units. The consumable may be a device as disclosed in International Patent Application No. PCT / US2024 / 036808, U.S. patent application 18 / 763,854, and U.S. Provisional Patent Application Serial No. 63 / 601 ,654, and U.S. Patent Application Attorney Docket No. ADDV-145 Titled “Sample Analysis Device and Methods” filed on November 8, 2024, and International Patent Application Attorney Docket No. ADDV-127WO2Titled “Two-Dimensional Matrix Droplet Array” filed on November 8, 2024 each of which are specifically incorporated by reference herein in their entirety.Scaling or combining assay processing units:

[0137] Embodiments of assay processing units, such as assay processing unit 300, are configured to be connected to one or more different automated assay processing units. By connected to different automated assay processing units, it is meant that assay processing units of the present disclosure are configured to operably connected to one or more instances of identical or substantially similar assay processing units. By operably connected, it is meant that such collection of interconnected assay processing units can operate together to perform a plurality of assays simultaneously or substantially simultaneously. Further, by operably connected, it is meant that such collection of interconnected assay processing units can share one or more resources among themselves, such as consumable resources, such as samples, buffers, reagents or the like, or one or more fixed resources, such as sample processing components or aspiration and dispense components or mixing components or optics components, or aspects thereof.

[0138] The plurality of operably connected assay processing components may be arranged in any convenient or desirable pattern. The plurality of assay processing components may be connected substantially in parallel or substantially in series. For example, in some cases, the one or more different automated assay processing units are adjacent to the automated assay processing unit such that the one or more automated assay processing units are parallel to the automated assay processing unit. For example, in other cases, the one or more different automated assay processing units are in series to the automated assay processing unit such that the one or more automated assay processing units are connected to the automated assay processing unit at the end closest to the first position or the third position.

[0139] Similarly, the plurality of operably connected assay processing units may be stacked in any convenient or desirable pattern. For example, in some cases, the one or more different automated assay processing units are vertically offset from the automated assay processing unit such that the one or more automated assay processing units are stacked above or below the automated assay processing unit. Various combinations of arrangements of operably connected assay processing units are also contemplated, such as where the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.

[0140] Arrangement 390-1 is a combination of two assay processing units, such as assay processing unit 300, operably connected in parallel. Arrangement 390-2 is a combination of tenassay processing units, such as assay processing unit 300, operably connected in parallel (two rows) and series (five columns). Arrangement 390-3 is a combination of 30 assay processing units, such as assay processing unit 300, operably connected in parallel (three rows) and series (ten columns). Arrangement 390-4 is a combination of 30 assay processing units, such as assay processing unit 300, operably connected in parallel (three rows) and series (ten columns).AUTOMATED MEDIUM-C PACITY ASSAY PROCESSING UNIT (50 TPH)

[0141] In some cases, embodiments of the present disclosure are automated medium-capacity assay processing units. By “automated medium-capacity assay processing units,” it is meant, for example, assay processing units configured to perform approximately 50 tests per hour, i.e., a 50 TPH unit. By tests, it is meant performing an assay, e.g., from an initial state with an unprocessed sample to a final state, in which assay results are obtained. Representative tests are illustrated in FIG. 1 and FIGS. 2A-2B. By performing approximately 50 TPH, it is meant that embodiments that are automated medium-capacity assay processing units are configured to perform typically 50 or more TPH, such as 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40, about 41, about 42, about 43, about 44, about 45, about 46, about 47, about 48, about 49, about 50, about 51, about 52, about 53, about 54, about 55, about 56, about 57, about 58, about 59, about 60 or more TPH. While embodiments that are automated mediumcapacity assay processing units are configured to perform up to 50 TPH, they need not always be used to run at maximum capacity, and instead may be used to perform 1 TPH or fewer.

[0142] FIG. 3S depicts automated medium-capacity assay processing unit 300A according to an embodiment. Automated medium-capacity assay processing unit 300A is a 50 TPH unit, and its configuration and relative size are shown relative to a standard assay processing unit 300 and an automated high-capacity assay processing unit 300B. FIG. 3U depicts automated mediumcapacity assay processing unit 300A according to an embodiment. In order to achieve a throughput of 50 TPH, in embodiments, automated medium-capacity assay processing unit 300A comprises: a plurality of stations that are dedicated to wash or conjugate mixing (i.e., six stations that are configured to perform similar functions as the second module, as described herein in connection with embodiments of a standard assay processing unit), such as six stations; a plurality of stations that are dedicated to fluid addition and / or mixing (i.e., three stations that are configured to perform similar functions as the third module comprising a mixing component, as described herein in connection with embodiments of a standard assay processing unit), such as three stations; and a plurality of stations that are dedicated to imaging (i.e., three stations that are configured to perform similar functions as the fourth module comprising an optics component, as describedherein in connection with embodiments of a standard assay processing unit), such as three stations. The usage of specified stations provides the benefit of processing multiple tests, i.e., assays, on multiple consumables simultaneous which not only increases the throughput of the automated assay processing units but also increases and / or maximizes resource utilization on the mediumcapacity assay processing unit. Embodiments of automated medium-capacity assay processing units further comprise a consumable mover configured to move one or more consumables between such stations. In embodiments, the consumable mover is configured to perform similar functions as the consumable holder, as described herein in connection with embodiments of a standard assay processing unit. The consumable mover may be a variety of movement mechanisms including, without limitation, a conveyer belt system, a robotic arm, a moveable stage or nest, a mechanism that engages with all or a portion of the consumable, etc.

[0143] Embodiments of automated medium-capacity assay processing unit may be configured to comprise a plurality of consumable holders such that automated medium-capacity assay processing units can hold a plurality of consumables at the same time. In embodiments, automated medium-capacity assay processing units are configured such that such consumables may be present in any of the plurality (such as six) of dedicated wash or conjugate mixing stations, any of the plurality (such as three) of dedicated fluid addition and / or mixing stations or any of the plurality (such as three) of dedicated imaging stations at the same time. That is, each of the dedicated wash or conjugate mixing stations, the dedicated fluid addition and / or mixing stations and the dedicated imaging stations are configured to operate on or manipulate more than one of the consumables present in the unit at substantially the same time. That is, each of the dedicated wash or conjugate mixing stations, the dedicated fluid addition and / or mixing stations, and the dedicated imaging stations are configured to operate substantially independently of each other such that the automated medium-capacity assay processing unit can process a plurality of consumables at the same time or otherwise achieve greater throughput, as compared with a standard assay processing unit 300.

[0144] In embodiments, a plurality of automated medium-capacity assay processing units may be operably connected with each other in parallel such that such collection of automated mediumcapacity assay processing units, together, achieve even greater through put, such as 150 TPH, when three automated medium-capacity assay processing units are operably connected 391-2, or 300 TPH, when six automated medium-capacity assay processing units are operably connected 391-3. Because each automated medium capacity assay processing unit is able to perform a specific assay or a collection of specific assays independently of other automated assay processing units, the assays may be configured to have: different durations of specific assay steps, different numbers of steps performed in a given assay, the ability to have each automated medium capacityassay processing unit to perform different assays, etc. The combination of multiple automated assay processing units also allows for an ease of maintenance because any given assay processing unit may be deactivated and repaired or maintained without the need to shutdown, turnoff, or alter the activity the other medium capacity automated processing units that are not undergoing maintenance.

[0145] Aspects of automated medium-capacity assay processing units comprise various modules and components that are described herein in connection with embodiments of standard assay processing units. Descriptions of such modules or components are not duplicated in connection with automated medium-capacity assay processing units.

[0146] Embodiments of automated medium-capacity assay processing units further comprise a degassing component. Any convenient degassing unit may be employed. In some embodiments, the degassing component comprises a vacuum component to remove air from one or more reagents present in the consumable. In some embodiments, the degassing component comprises a cooling component to cool one or more reagents present in the consumable thereby reducing the air present in one or more reagents in the consumable.Consumable mover:

[0147] The consumable mover may be a variety of movement mechanisms including, without limitation, a conveyer system, a robotic arm or gripper, a moveable stage or nest, a mechanism that engages with all or a portion of the consumable, etc. In embodiments of the automated medium-capacity assay processing unit, the consumable mover comprises a conveyer system. In some cases, such conveyer system is operably coupled to the fluid addition stations, wash stations and imaging stations. In some cases, the consumable mover is configured to move a consumable from a fluid addition station to a wash station and / or from a wash station to an imaging station. In embodiments, the conveyer system comprises a consumable holder that is capable of being moved between each station; e.g., between any fluid addition station, any wash station and any imaging station. In embodiments, the consumable mover comprises a griping component that is capable of gripping a consumable and moving the consumable between stations. In some embodiments, the consumable mover moves all consumables in the automated medium capacity assay processing unit at once. In some embodiments, the consumable mover moves only a portion of the consumables in the automated medium capacity assay processing unit at once. In some embodiments, the consumable mover moves consumables in the automated medium capacity assay processing unit selectively such that only intended consumables are moved and unintended consumables are not moved. By “intended” it is meant that the consumable is intentionally movedbecause, for example, an assay process has been completed, such as an assay process step disclosed in FIG. 1.Heating elements:

[0148] In embodiments of the automated medium-capacity assay processing unit, each of the plurality of fluid addition stations comprise one or more heating elements. In some cases, such heating elements are configured to heat one or more consumables to a first temperature when the one or more consumables are present in a fluid addition station. Any convenient heating element, such as resistive heating elements may be applied. In some cases, the first temperature is a temperature that is 0 C or greater, such as 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the first temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25- 30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C, about 55-60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than about 100 C.

[0149] In embodiments of the automated medium-capacity assay processing unit, each of the plurality of wash stations comprise one or more heating elements. In some cases, such heating elements are configured to heat one or more consumables to a second temperature when the one or more consumables are present in a fluid addition station. Any convenient heating element, such as resistive heating elements may be applied. In some cases, the second temperature is a temperature that is 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the second temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C, about 55- 60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than 100 C.

[0150] In embodiments, the first temperature may differ from the second temperature, i.e., such that consumable, or aspects thereof, is heated to a first temperature when present in fluid addition stations and heated to a different temperature when present in washing stations. First temperature may be greater than second temperature by any convenient amount, such as by about 0 C, about 5 C, about 10 C, about 15 C, about 20 C or greater. First temperature may be less than secondtemperature by any convenient amount such as by about 0 C, about 5 C, about 10 C, about 15 C, about 20 C or greater. In other embodiments, the first temperature may be the same as the second temperature, i.e., such that consumable, or aspects thereof, is heated to a first temperature when present in fluid addition stations and heated to the same temperature in washing stations.

[0151] In embodiments, the two or more fluid addition stations comprises a first temperature sensor. In embodiments, two or more fluid addition stations may be configured such that the first temperature sensor senses a temperature, e.g., of consumable, or aspects of consumable, or the ambient temperature in the fluid addition stations. In embodiments, two or more wash stations comprise a second temperature sensor. In embodiments, two or more wash stations may be configured such that the second temperature sensor senses a temperature, e.g., of consumable, or aspects of consumable, or the ambient temperature in washing stations. In embodiments, any convenient temperature sensor, such as any convenient commercially available temperature sensor, such as, for example, one or more ambient temperature sensors, may be utilized for the first and second temperature sensors.Fluid addition stations:

[0152] In embodiments of the automated medium-capacity assay processing unit, the two or more fluid addition stations comprise one or more reagent dispensers. In some cases, the reagent dispensers comprise one or more reagent pumps. In some cases, the reagent pumps are fluidically connected to one or more reagent nozzles. In embodiments, the two or more fluid addition stations share the one or more reagent dispensers. In other cases, each fluid addition station comprises the one or more reagent dispensers. It is contemplated that any convenient fluid, including any convenient reagent, capable of being dispensed through a reagent pump may be dispensed. The one or more reagent dispensers may dispense a single type of reagent or may dispense multiple types of reagents. In some embodiments, the one or more reagent dispensers are two or more, three or more, four or more, five or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more reagent dispensers.

[0153] In embodiments, the two or more fluid addition stations comprise a hydrophilic liquid dispenser. In some cases, the hydrophilic liquid dispenser comprises a hydrophilic liquid nozzle. In some cases, the hydrophilic liquid nozzle is fluidically connected to a hydrophilic liquid pump. In embodiments, the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle. In some cases, the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle when the consumable is in fluid addition stations; i.e., wherein the consumable holder positions a consumable, or aspects thereof, such that consumable can receive liquid from the nozzle.

[0154] In embodiments, the two or more fluid addition stations comprise a mixing component. In some cases, the two or more fluid addition stations share the mixing component; i.e., a plurality of fluid addition stations share a single mixing component. In other cases, each fluid addition station comprises a dedicated mixing component. In embodiments, the mixing component comprises a mixing motor and an engagement arm. In some cases, the engagement arm is configured to engage with a consumable when the consumable is present in a fluid addition station. In some cases, the engagement arm engages with a portion of the consumable. In other cases, the engagement arm engages with the entire consumable. In embodiments, the mixing component comprises a ballistic mixing device. In some cases, the ballistic mixing device is a pipettor.Wash stations:

[0155] In embodiments of the automated medium-capacity assay processing unit, the two or more wash stations comprise a first magnet. In some cases, the first magnet is a retractable magnet. In some cases, the first magnet is positioned beneath a consumable. In some cases, the first magnet is positioned beneath a consumable when the consumable is present in one of the two or more wash stations. In some cases, wherein the magnet is a retractable magnet, the first magnet is configured to be proximate to the consumable in an un-retracted state. In some cases, wherein the magnet is a retractable magnet, the first magnet is configured to be distal to the consumable in a retracted state. In embodiments, wherein the magnet is a retractable magnet, the first retractable magnet is encased in a sleeve in the retracted state wherein the sleeve shields the first retractable magnet thereby reducing the local magnetic field.

[0156] In some cases, the two or more wash stations share a first retractable magnet. In other cases, each wash station comprises its own dedicated first retractable magnet.

[0157] In embodiments, the two or more wash stations comprise one or more heating elements configured to heat one or more consumables to a second temperature when the one or more consumables are present in the two or more wash stations. Any convenient heating element, such as resistive heating elements, may be applied. In some cases, the second temperature is a temperature that is about 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the second temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C, about 55-60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than about 100 C.

[0158] In embodiments, the two or more wash stations comprise a second magnet. In some cases, the second magnet is a second retractable magnet. In some cases, the second retractable magnet is positioned above a consumable when the consumable is in a wash station of the two or more wash stations. In some cases, wherein the magnet is a retractable magnet, the second retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state.

[0159] In embodiments, the second magnet is positioned in a position that is horizontally offset from the first magnet. In embodiments, wherein the magnets are retractable magnets, the first retractable magnet and the second retractable magnet are configured to not be in the un-retracted state at the same time. That is, the first and second retractable magnets are interlocked so that they cannot both engage the consumable at the same time. The first magnet and the second magnet may be un-retracted and retracted in an alternating fashion such that alternating magnet fields may be applied to the consumable. The consumable and consumable holder may be moved between the first magnet and the second magnet in an alternating fashion, e.g., from the first magnet to the second magnet in a horizontal position, in order to alternate the magnetic fields on the consumable.

[0160] In embodiments, the two or more wash stations comprise an aspiration device. In some cases, the aspiration devices comprises an aspiration nozzle. In some cases, the aspiration nozzle is fluidically connected to an aspiration pump. In embodiments, the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle when the hydrophilic liquid is present in the consumable. In some cases, the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle when a consumable is present in the two or more wash stations. In some cases, the two or more wash stations share the aspiration device. In other cases, each wash station comprises an aspiration device. That is, each wash station comprises a dedicated aspiration device.

[0161] In embodiments, the two or more wash stations comprise a hydrophobic liquid dispenser. In some cases, the hydrophobic liquid dispenser comprises a hydrophobic liquid nozzle. In some cases, the hydrophobic liquid nozzle is fluidically connected to a hydrophobic liquid pump. In embodiments, the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle. In some cases, the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle when a consumable is present in the two or more wash stations. In embodiments, the aspiration pump is configured to aspirate the hydrophilic liquid. In some cases, the hydrophobic liquid pump is configured to dispense the hydrophobic liquid. In some cases, the aspiration pump is configured to aspirate the hydrophilic liquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously. In some cases, the aspiration pump is configured to aspirate the hydrophilicliquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously when a consumable is present in the two or more wash stations.

[0162] In embodiments, the two or more wash stations share the hydrophobic liquid dispenser. In some cases, each wash station comprises a dedicated hydrophobic liquid dispenser. In embodiments, the hydrophobic liquid dispenser is a first hydrophobic liquid dispenser of a plurality of hydrophobic liquid dispensers, and each wash station comprises a dedicated hydrophobic liquid dispenser of the plurality of hydrophobic liquid dispensers.

[0163] In embodiments, the two or more wash stations comprise a mixing component. In some cases, the two or more wash stations share the mixing component; i.e., a plurality of wash stations share a single mixing component. In other cases, each wash station comprises a dedicated mixing component. In embodiments, the mixing component comprises a mixing motor and an engagement arm. The mixing motor may be a range of different motors, including, without limitation, a stepper motor, a brushless DC motor, a solenoid, etc. In some embodiments, the mixing motor is a voice coil. In some cases, the engagement arm is configured to engage with a consumable when the consumable is present in a wash station. In some cases, the engagement arm engages with a portion of the consumable. In other cases, the engagement arm engages with the entire consumable.

[0164] In embodiments, the engagement arm is attached to a chamber in the shape of a cube, rectangle, or cylinder comprising a diaphragm on the interior and an opening at one end with a suction cup or a gasket. The suction cup or the gasket contacts a portion of the consumable. The diaphragm may be moved or oscillated by a deflection amount that compresses the air in the chamber. The compression in the air results in movement in the fluid consumable thereby mixing the fluid. The chamber comprises one or more openings that allow pressure relief in the chamber when the diaphragm is moving or oscillating. The chamber may be according to the pressure mixing device disclosed in US Patent Application No. 18 / 763,854.

[0165] The diaphragm of the sample mixing device may be made of a range of different materials including, without limitation, piezoelectric material, metal, a compliant material that deforms under heat or pressure, etc. The diaphragm of the sample mixing device may be moved or oscillated in a number of different ways. In some embodiments, the diaphragm is made of a piezoelectric material. In some embodiments, the piezoelectric material is moved or oscillated by applying an electric current to the piezoelectric material. In some embodiments, the diaphragm is metal. In some embodiments, the metal is moved or oscillated by turning a magnetic field on and off. In some embodiments, the diaphragm is made of a compliant material that deforms under heat or pressure. In some embodiments, the compliant material is moved by apply air pressure to the compliant material. In some embodiments, the compliant material is moved by applying heat tothe compliant material. In some embodiments, the compliant material is moved by applying acoustic waves to the compliant material. In some embodiments, the compliant material is moved by applying a mechanical force to the compliant material.

[0166] In embodiments, the mixing component comprises a ballistic mixing device. In some cases, the ballistic mixing device is a pipettor.Imaging Stations:

[0167] In embodiments of the automated medium-capacity assay processing unit, the two or more imaging stations comprise an imaging component. In some cases, the imaging component is positioned beneath a consumable when the consumable present in the two or more imaging stations. In some cases, the imaging component is positioned beneath a consumable when the consumable is present in one of two or more imaging stations. That is, when consumable holder is positioned in one of two or more imaging stations, an imaging component is aligned with consumable or one or more wells, well arrays, or partitions of consumable. In some cases, the imaging component is positioned above a consumable when the consumable present in the two or more imaging stations. In some cases, the imaging component is positioned above a consumable when the consumable is present in one of two or more imaging stations. That is, when consumable holder is positioned in one of two or more imaging stations, an imaging component is aligned with consumable or one or more wells, well arrays, or partitions of consumable.

[0168] In embodiments, the imaging component comprises an illumination component. The illumination component may be configured to facilitate imaging consumable by illuminating consumable or one or more portions of consumable. In some cases, the illumination component comprises one or more lighting elements. For example, the illumination component may comprise one or more LEDs. In some cases, the illumination component is enclosed or encased or present within enclosed illumination chamber. In some cases, enclosed illumination chamber comprises opening or window or aperture. In embodiments, opening or window or aperture is positioned beneath consumable holder when consumable holder is present in one of the two or more imaging stations. In embodiments, opening or window or aperture is positioned above the consumable holder when the consumable holder is present in one of the two or more imaging stations. That is, when consumable holder is present in one of the two or more imaging stations, such opening or window or aperture is aligned with consumable or one or more wells, well arrays, or partitions of consumable, such that opening or window or aperture allows consumable or aspects thereof to be illuminated and / or imaged by illumination component, e.g., LEDs, and / or imaging component. In other words, system is configured such that when consumable is positioned in one of the two or more imaging stations, such aperture of imaging component aligns with consumable, or an aspectof consumable, such as a well or partition thereof, such that consumable can be illuminated and / or imaged through aperture.

[0169] In embodiments, the one or more LEDs are in proximity to one or more temperature sensing elements, such as, for example, thermistors. In embodiments, the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs. In embodiments, the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a pattern, such as a circular pattern. In embodiments, the one or more thermistors are two or more, or three or more thermistors. In embodiments, the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs. In some cases, the one or more LEDs are present on a printer circuit board (PCB). In embodiments, the PCB maintains one or more of the LEDs, the enclosed illumination chamber, and the consumable holder, when the consumable holder is located at an imaging station at a third temperature. In some cases, the third temperature is a temperature that is about 100 C or lower, such as about 90 C, about 85 C, about 80 C, about 75 C, about 60 C, about 55 C, about 50 C, about 45 C, about 40 C or less. In some embodiments, the PCB is configured to maintain the LEDs between about 65 and 45 C.

[0170] In embodiments, PCB comprises a core. In embodiments, the core may comprise a material selected to improve heat dissipation from PCB or from resistive elements present on PCB . Materials that improve heat dissipation include, without limitation, silver, copper, gold, aluminum, iron, nickel, brass, tungsten, zinc, alloys and combinations thereof, etc. In embodiments, PCB comprises an aluminum core.

[0171] In embodiments, the imaging component comprises a clamping component. In some cases, the clamping component engages with a consumable and provides a downward clamping force on the consumable when the consumable is present in the two or more imaging stations. In embodiments, the imaging component comprises a clamping component configured to engage the consumable holder or to engage the consumable present in the consumable holder. The clamping component may be further configured to engage the consumable holder or the consumable by applying a downward clamping force to the consumable holder or the consumable. In some cases, clamping component is configured to engage consumable holder or consumable when consumable holder is present in the imaging station. In embodiments, the clamping component may be configured to hold the consumable in a fixed position such that aspects of the imaging component, such as one or more aspects of the imaging component, can be engaged or controlled to collect one or more images of the consumable or the contents thereof. In embodiments, the clamping component may be configured to hold the consumable substantially in a fixed position for a period of time during which the imaging station, or imaging component thereof, images one or moreaspects of the consumable, such as one or more sections of the consumable over time. In embodiments, the clamping component may comprise one or more constituent components, such as, for example, one or more springs or one or more clamps or one or more motors or one or more cams. In embodiments, the clamping component depresses (i.e., pushes down on) the consumable holder or consumable when the consumable holder or the consumable physically contacts aspects of the imaging station, such as the imaging component thereof. In embodiments, when the consumable holder is positioned in an imaging station, the clamping component is aligned with the consumable such that the clamping component can apply a downward force on the consumable. In other words, the automated medium capacity assay processing unit is configured such that when the consumable is positioned in the imaging station, the clamping component of the imaging component aligns with the consumable, or an aspect of the consumable, such that the consumable can held in a fixed position, e.g., a reference position abutting an alignment datum pin by the clamping component.

[0172] The imaging stations may comprise any of the imaging components described above.Scaling or combining automated medium-capacity assay processing units:

[0173] In embodiments, automated medium-capacity assay processing units are configured to be capable of being connected to one or more different automated assay processing units, such as one or more automated medium-capacity assay processing units. In some cases, the one or more different automated medium-capacity assay processing units are adjacent to the automated medium-capacity assay processing unit such that the one or more automated medium-capacity assay processing units are parallel to the automated medium-capacity assay processing unit. In other cases, the one or more different automated medium-capacity assay processing units are in series to the automated medium-capacity assay processing unit such that the one or more automated medium-capacity assay processing units are connected to the automated mediumcapacity assay processing unit at an end, i.e., end-to-end or in serial. In still other cases, the one or more different automated medium-capacity assay processing units are vertically offset from the automated medium-capacity assay processing unit such that the one or more automated mediumcapacity assay processing units are stacked above or below the automated medium-capacity assay processing unit. In further cases, the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.Consumable:

[0174] Embodiments of automated medium-capacity assay processing units are configured to accept and operate on a plurality of consumables simultaneously. Embodiments of automated medium-capacity assay processing units are configured to accept and operate on consumables that are identical to consumables utilized in embodiments of standard assay processing units or automated high-capacity assay processing units.AUTOM TED HIGH-C P CITY ASSAY PROCESSING UNITS (150 TPH)

[0175] In some cases, embodiments of the present disclosure are automated high-capacity assay processing units. By “automated high-capacity assay processing units,” it is meant, for example, assay processing units configured to perform approximately 150 tests per hour; i.e., a 150 TPH unit. By “test”, it is meant performing an assay, e.g., from an initial state with an unprocessed sample to a final state, in which assay results are obtained. Representative tests are illustrated in FIG. 1 and FIGS. 2A-2B. By performing approximately 150 TPH, it is meant that embodiments that are automated medium-capacity assay processing units are configured to perform typically 150 or more TPH, such as about 140, about 141, about 142, about 143, about 144, about 145, about 146, about 147, about 148, about 149, about 150, about 151, about 152, about 153, about 154, about 155, about 156, about 157, about 158, about 159, about 160 or more TPH. While embodiments that are automated high-capacity assay processing units are configured to perform up to 150 TPH, they need not always be used to run at maximum capacity, and instead may be used to perform 1 TPH or fewer.

[0176] FIG. 3S depicts automated high-capacity assay processing unit 300B according to an embodiment. Automated high-capacity assay processing unit 300B is a 150 TPH unit, and its configuration and relative size are shown relative to a standard assay processing unit 300 and automated medium-capacity assay processing unit 300A. FIG. 3V depicts automated high- capacity assay processing unit 300B according to an embodiment. In order to achieve a throughput of 150 TPH, in embodiments, automated high-capacity assay processing unit 300B comprises; a plurality of stations that are dedicated to washing, mixing and seeding (e.g., two carousels stations, each with 18 stations, that are configured to perform similar functions as the second module, as described herein in connection with embodiments of a standard assay processing unit), such as 36 stations; a plurality of stations that are dedicated to fluid addition and / or mixing (e.g., eight stations that are configured to perform similar functions as the third module comprising a mixing component, as described herein in connection with embodiments of a standard assay processing unit), such as eight stations; and a plurality of stations that are dedicated to imaging (e.g., eight stations that are configured to perform similar functions as the fourth module comprising an optics component, as described herein in connection withembodiments of a standard assay processing unit), such as eight stations. Embodiments of automated high -capacity assay processing units further comprise a consumable mover configured to move one or more consumables between such stations. In embodiments, the consumable mover is configured to perform similar functions as the consumable holder, as described herein in connection with embodiments of a standard assay processing unit. In embodiments, the consumable mover comprises one or more carousels. In embodiments, the consumable mover comprises one or more carousel interfaces for loading onto and unloading from a carousel.

[0177] Embodiments of automated high-capacity assay processing unit may be configured to comprise a plurality of consumable holders such that automated high-capacity assay processing units can hold a plurality of consumables at the same time. In embodiments, automated high- capacity assay processing units are configured such that such consumables may be present in any of the plurality (such as 36) of dedicated wash or conjugate mixing stations, any of the plurality (such as eight) of dedicated fluid addition and / or mixing stations or any of the plurality (such as eight) of dedicated imaging stations at the same time. That is, each of the dedicated wash or conjugate mixing stations, the dedicated fluid addition and / or mixing stations and the dedicated imaging stations are configured to operate on or manipulate more than one of the consumables present in the unit at substantially the same time. That is, each of the dedicated wash or conjugate mixing stations, the dedicated fluid addition and / or mixing stations and the dedicated imaging stations are configured to operate substantially independently of each other such that the automated high-capacity assay processing unit can process a plurality of consumables at the same time or otherwise achieve greater throughput, as compared with a standard assay processing unit 300.

[0178] In embodiments, a plurality of automated high-capacity assay processing units may be operably connected with each other in parallel such that such collection of automated high- capacity assay processing units, together, achieve even greater throughput, such as 300 TPH, when two automated high-capacity assay processing units are operably connected 391-3.

[0179] Aspects of automated high-capacity assay processing units comprise various modules and components that are described herein in connection with embodiments of standard assay processing units and / or automated medium-capacity assay processing units. Descriptions of such modules or components are not duplicated in connection with automated high-capacity assay processing units.

[0180] In embodiments, automated high-capacity assay processing unit, comprise four or more fluid addition stations, two or more wash stations, wherein each wash station comprises a first carousel encircled by a second carousel, four or more imaging stations, and a consumable mover.Details regarding such components employed in high-capacity assay processing units are provided herein.Consumable mover:

[0181] The consumable mover may be a variety of movement mechanisms including, without limitation, a conveyer system, a robotic arm or gripper, a moveable stage or nest, a mechanism that engages with all or a portion of the consumable, etc. In embodiments of an automated high- capacity assay processing unit, the consumable mover comprises a conveyer system. In some cases, the conveyer system is operably coupled to each of the fluid addition stations, wash stations, and imaging stations. That is, the conveyer system is configured to translate among the fluid addition stations, wash stations, and imaging stations. In some cases, the conveyer system is configured to translate among each fluid addition station, each wash station, and each imaging station. In other cases, the conveyer system is configured to translate among a subset of the fluid addition stations, a subset of the wash stations, and a subset of the imaging stations. In embodiments, the consumable mover is configured to move a consumable. In embodiments, the consumable mover is configured to move a consumable from a fluid addition station of the plurality of fluid addition stations to a wash station of the plurality of wash stations, and from the wash station to an imaging station of the plurality of imaging stations.

[0182] In embodiments, the conveyer system comprises a consumable holder, i.e., for holding a consumable. In embodiments, the consumable holder of the conveyer system moves between each station, or a subset of stations, of the unit. In embodiments, the consumable mover comprises a griping component. Such griping component may be configured such that it is capable of gripping a consumable and moving the consumable between each station, or a subset of stations, of the unit. Any convenient gripping component may be employed.Fluid addition stations:

[0183] In embodiments, the four or more fluid addition stations comprise one or more heating elements. Such heating elements may be configured to heat one or more consumables, i.e., consumables or aspects thereof, present in the unit. In embodiments, heating elements may be configured to heat a consumable to a first temperature. The first temperature may be any convenient temperature, such as a temperature selected to facilitate one or more steps of loading, unloading, IR1, IR2, mixing and dispensing, ER or imaging-related steps. In some cases, the first temperature is a temperature that is 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about95 C, about 100 C or greater. In some embodiments, the first temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about SO- 55 C, about 55-60 C, about 60-65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than about 100 C. In embodiments, heating elements may be configured to heat one or more consumables when the one or more consumables are present in the four or more fluid addition stations. Any convenient heating elements may be applied, as such are described herein, for example resistive heating elements.

[0184] In embodiments, the four or more fluid addition stations comprise one or more reagent dispensers. Reagent dispensers may be any convenient dispenser capable of dispensing reagent into one or more aspects of a consumable. In embodiments, reagent dispensers comprise one or more reagent pumps fluidically connected to one or more reagent nozzles. In some cases, the four or more fluid addition stations share the one or more reagent dispensers. In other cases, each fluid addition station comprises the one or more reagent dispensers. That is, each fluid addition station comprises a dedicated reagent dispenser. The one or more reagent dispensers may dispense a single type of reagent or may dispense multiple types of reagents. In some embodiments, the one or more reagent dispensers are two or more, three or more, four or more, five or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more reagent dispensers. In some embodiments, the number of reagent dispensers is equivalent to the number of reagents used in an assay.

[0185] In embodiments, the four or more fluid addition stations comprise a hydrophilic liquid dispenser. Hydrophilic liquid dispensers may be any convenient dispenser capable of dispensing hydrophilic liquid into one or more aspects of a consumable. In embodiments, hydrophilic liquid dispenses comprise a hydrophilic liquid nozzle fluidically connected to a hydrophilic liquid pump. In embodiments, the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle. In embodiments, the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle when the consumable holder, and consumable gripped therein, is located in a fluid addition station.Mixing component:

[0186] In embodiments, the four or more fluid addition stations comprise a mixing component. For example, in some cases, each of the four or more fluid addition stations comprises a dedicated mixing component. In other cases, the four or more fluid addition stations share the mixing component, the mixing component comprises a mixing motor and an engagement arm that is configured to engage with a consumable when the consumable is present in the four or more fluidaddition stations. In embodiments, the mixing component comprises a mixing motor and an engagement arm. The engagement arm is configured to engage with a consumable when the consumable is present in a fluid addition station of the four or more fluid addition stations. In some cases, the engagement arm engages with a portion of the consumable. In other cases, the engagement arm engages with the entire consumable.

[0187] In embodiments, the two or more wash stations comprise a mixing component. For example, in some cases, each of the two or more wash stations comprises a dedicated mixing component. In other cases, the two or more wash stations share the mixing component. The mixing component comprises a mixing motor and an engagement arm that is configured to engage with a consumable when the consumable is present in the two or more wash stations. In embodiments, the mixing component comprises a mixing motor and an engagement arm. The engagement arm is configured to engage with a consumable when the consumable is present in a wash station of the two or more wash stations. In some cases, the engagement arm engages with a portion of the consumable. In other cases, the engagement arm engages with the entire consumable.

[0188] In embodiments, the mixing component comprises a ballistic mixing device. For example, in some cases, the ballistic mixing device is a pipettor. The mixing component may be any of the mixing components described above.First Magnet:

[0189] In embodiments, the two or more wash stations comprise a first magnet. In embodiments, the first magnet is a first retractable magnet. In embodiments, the first magnet is positioned beneath a consumable when the consumable is present in one of the two or more wash stations. In some cases, the first retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state. In some embodiments in which the first magnet is a first retractable magnet, the first retractable magnet is encased in a sleeve in the retracted state wherein the sleeve shields the first retractable magnet thereby reducing the local magnetic field.

[0190] In some cases, the two or more wash stations share the first retractable magnet. In other cases, each wash station comprises the first retractable magnet. That is, each wash station comprises a dedicated retractable magnet. Some embodiments comprise a plurality of first retractable magnets.Wash station heating elements:

[0191] In embodiments, the two or more wash stations comprise one or more heating elements. In embodiments, such heating elements are configured to heat one or more consumables or aspects thereof. In embodiments, such heating elements are configured to heat one or more consumables or aspects thereof, to a second temperature when the one or more consumables are present in the two or more wash stations. The second temperature may be any convenient temperature, such as a temperature selected to facilitate one or more steps of loading, unloading, 1R1, IR2, mixing and dispensing, ER or imaging-related steps. In some cases, the second temperature is a temperature that is 0 C or greater, such as about 0 C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some embodiments, the second temperature is a temperature range. For instance, the range may be about 0-5 C, about 5-10 C, about 10-15 C, about 15-20 C, about 20-25 C, about 25-30 C, about 30-35 C, about 35-40 C, about 40-45 C, about 45-50 C, about 50-55 C, about 55-60 C, about 60- 65 C, about 65-70 C, about 70-75 C, about 75-80 C, about 80-85 C, about 85-90 C, about 90-95 C, about 95-100 C, or greater than about 100 C.Second magnet:

[0192] Embodiments of the high-capacity assay processing unit comprise two or more wash stations comprising a second magnet. In embodiments, the second magnet is a second retractable magnet. In embodiments, the second magnet is positioned above a consumable when the consumable is in the two or more wash stations. In some embodiments in which the second magnet is a second retractable magnet, the second retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state. In embodiments, the second retractable magnet is positioned in a position that is horizontally offset from the first retractable magnet. In embodiments, the first retractable magnet and the second retractable magnet are configured to not be in the un-retracted state at the same time. That is, first and second retractable magnets are interlocked to prevent them both from being extended in an un-retracted state simultaneously. Embodiments comprise a plurality of second retractable magnets. The first magnet and the second magnet may be un-retracted and retracted in an alternating fashion such that alternating magnet fields may be applied to the consumable. The consumable and consumable holder may be moved between the first magnet and the second magnet in an alternating fashion, e.g., from the first magnet to the second magnet in a horizontal position, in order to alternate the magnetic fields on the consumable. The first magnet and the second magnet may be moved or have a set of programed movements based on the position of the carousel described below. For instance, when the carousel rotates, the first magnet and the secondmagnet may be moved or have a set of programed movements and actions that are initiated once rotation commences or ceases. In some embodiments, the first magnet and the second magnet only move when a consumable is detected in a specific location within a carousel.Wash station aspiration device;

[0193] tn embodiments, the two or more wash stations comprise an aspiration device. In embodiments, the aspiration device comprises an aspiration nozzle fluidically connected to an aspiration pump. In some cases, the aspiration pump is configured to aspirate a hydrophilic liquid. In some cases, the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle when a consumable is present in the two or more wash stations. In some instances, the two or more wash stations share the aspiration device. In other instances, each wash station comprises the aspiration device. That is, in certain instances, each wash station comprises a dedicated aspiration device. The aspiration device may be moved or have a set of programmed movements and actions based on the position of a carousel described below. For instance, when the carousel rotates, the aspiration device may be moved or have a set of programmed movements and actions that are initiated once rotation of a carousel commences or ceases. In some embodiments, the aspiration device aspirates a hydrophilic liquid when the consumable is present in a specific location within a carousel.Hydrophobic liquid dispenser;

[0194] In embodiments, the two or more wash stations comprise a hydrophobic liquid dispenser. In embodiments, the hydrophobic liquid dispenser comprises a hydrophobic liquid nozzle fluidically connected to a hydrophobic liquid pump. In embodiments, the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle. In embodiments, the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle when a consumable is present in one of the two or more wash stations. The hydrophobic liquid dispenser may be moved or have a set of programmed movements and actions based on the position of a carousel described below. For instance, when the carousel rotates, the hydrophobic liquid dispenser may be moved or have a set of programmed movements and actions that are initiated once rotation of a carousel commences or ceases. In some embodiments, the hydrophobic liquid dispenser dispenses a hydrophobic liquid when the consumable is present in a specific location within a carousel.

[0195] Tn embodiments, the four or more fluid addition stations comprise a hydrophobic liquid dispenser. In embodiments, the hydrophobic liquid dispenser comprises a hydrophobic liquid nozzle fluidically connected to a hydrophobic liquid pump. In embodiments, the hydrophobicliquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle. In embodiments, the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle when a consumable is present in one of the four or more fluid addition stations. The hydrophobic liquid dispenser may be moved or have a set of programmed movements and actions based on the position of a carousel described below. For instance, when the carousel rotates, the hydrophobic liquid dispenser may be moved or have a set of programmed movements and actions that are initiated once rotation of a carousel commences or ceases. In some embodiments, the hydrophobic liquid dispenser dispenses a hydrophobic liquid when the consumable is present in a specific location within a carousel.

[0196] In embodiments, the aspiration pump is configured to aspirate the hydrophilic liquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously when a consumable is present in one of the two or more wash stations.

[0197] In some cases, the two or more wash stations share the hydrophobic liquid dispenser. In other cases, each wash station comprises the hydrophobic liquid dispenser. That is, each wash station comprises a dedicated hydrophobic liquid dispenser.

[0198] In some cases, the four or more fluid addition stations share the hydrophobic liquid dispenser. In other cases, each fluid addition station comprises the hydrophobic liquid dispenser. That is, each fluid addition station comprises a dedicated hydrophobic liquid dispenser.

[0199] When the four or more fluid addition stations comprise the hydrophilic liquid dispenser, the two or more wash stations do not comprise the hydrophilic liquid dispenser. When the two or more wash stations comprise the hydrophilic liquid dispenser, the four or more fluid addition stations do not comprise the hydrophilic liquid dispenser.Carousels:

[0200] In embodiments, the first carousel has a lower capacity for consumables than the second carousel. That is, in some cases, the first carousel is configured to receive a larger number of consumables than the second carousel, such as one or two or three or four or five or six or seven or eight or nine or ten or 20 or 30 or 40 or 50 or more consumables. The size of carousel may equate to the number of assay steps that are performed in the consumable. For instance, a larger outer carousel, i.e., the second carousel, may be used for a two-step assay whereas a smaller inner carousel, i.e., the first carousel, may be used for a one-step assay. By “two step assay” it is meant that two assay steps are performed that do not include washing steps. By “one step assay” it is meant that one assay step is performed that does not include washing steps.

[0201] In embodiments, the first carousel rotates in lockstep with the second carousel. That is, the first and second carousels are configured such that when one carousel rotates, the other isnecessarily also rotates, i.e., rotates by the same or similar angular displacement, for example. In other embodiments, the first carousel rotates out of sync with the second carousel. That is, the first and second carousels are independent or decoupled with respect to rotation, such that the first carousel can rotate while the second carousel does not rotate or rotates in a different direction or at a different velocity. Specific movements or actions of components in the wash stations, e.g., the first magnet and the second magnet, the hydrophobic liquid dispenser, the aspiration device, the mixing component; may be initiated when the carousel rotates or ceases rotation. Specific movements or actions of components in the wash station may not occur if a consumable is not in a specific location.Imaging Stations:

[0202] In embodiments of high-capacity assay processing units, the two or more imaging stations comprise an imaging component. In some cases, the imaging component is positioned beneath a consumable when the consumable present in one of the two or more imaging stations. That is, the imaging component is positioned in the unit such that it is beneath the consumable when the consumable is positioned in an imaging station. In some cases, the imaging component is positioned above a consumable when the consumable present in one of the two or more imaging stations. That is, the imaging component is positioned in the unit such that it is above the consumable when the consumable is positioned in an imaging station.

[0203] In embodiments, the imaging component comprises an illumination component comprising one or more lighting elements, such as one or more LEDs. In embodiments, the illumination component is encased in an enclosed illumination chamber having an opening or window or aperture positioned beneath each imaging station. In some cases, the one or more lighting elements, e.g., LEDs, are in proximity to one or more thermistors. In embodiments, the one or more lighting elements, e.g., LEDs, are two or more, three or more, four or more, five or more, or six or more lighting elements, e.g., LEDs. Such lighting elements, e.g., LEDS, may be arranged in a pattern, e.g., a circular pattern. In embodiments, the one or more thermistors are two or more, or three or more thermistors. In some cases, the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs. In some cases, the one or more lighting elements, e.g., LEDs, are present on a printed circuit board (PCB). In some cases, the PCB maintains the LEDs, the enclosed illumination chamber, and the consumable holder, when the consumable holder is at the third position, at a third temperature. The third temperature may be any convenient temperature, such as a temperature selected to facilitate one or more steps of loading, unloading, IR1, IR2, mixing and dispensing, ER or imaging-related steps. In some cases, the third temperature is a temperature that is 0 C or greater, such as about 0C, about 5 C, about 10 C, about 15 C, about 20 C, about 25 C, about 30 C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 C, about 90 C, about 95 C, about 100 C or greater. In some cases, the third temperature is a temperature that is 100 C or lower, such as about 90 C, about 85 C, about 80 C, about 75 C, about 60 C, about 55 C, about 50 C, about 45 C, about 40 C or less. In some embodiments, the PCB is configured to maintain the LEDs between about 65 and 45 C.

[0204] In embodiments, the PCB comprises a core, such as a metallic core, such as an aluminum core. The PCB may be any of the PCBs described above.

[0205] In embodiments, the imaging component comprises a clamping component. In some cases, the clamping component engages with a consumable and provides a downward clamping force on the consumable when the consumable is present in the two or more imaging stations. In embodiments, the imaging component comprises a clamping component configured to engage the consumable holder or to engage the consumable present in the consumable holder. The clamping component may be further configured to engage the consumable holder or the consumable by applying a downward clamping force to the consumable holder or the consumable. In some cases, clamping component is configured to engage consumable holder or consumable when consumable holder is present in the imaging station. In embodiments, the clamping component may be configured to hold the consumable in a fixed position such that aspects of the imaging component, such as one or more aspects of the imaging component, can be engaged or controlled to collect one or more images of the consumable or the contents thereof. In embodiments, the clamping component may be configured to hold the consumable substantially in a fixed position for a period of time during which the imaging station, or imaging component thereof, images one or more aspects of the consumable, such as one or more sections of the consumable over time. In embodiments, the clamping component may comprise one or more constituent components, such as, for example, one or more springs or one or more clamps or one or more motors or one or more cams. In embodiments, the clamping component depresses (i.e., pushes down on) the consumable holder or consumable when the consumable holder or the consumable physically contacts aspects of the imaging station, such as the imaging component thereof. In embodiments, when the consumable holder is positioned in an imaging station, the clamping component is aligned with the consumable such that the clamping component can apply a downward force on the consumable. In other words, the automated high capacity assay processing unit is configured such that when the consumable is positioned in the imaging station, the clamping component of the imaging component aligns with the consumable, or an aspect of the consumable, such that the consumable can held in a fixed position, e.g., a reference position abutting an alignment datum pin by the clamping component.

[0206] Embodiments of automated high-capacity assay processing units further comprise a degassing component. Any convenient degassing unit may be employed. In some embodiments, the degassing component comprises a vacuum component to remove air from one or more reagents present in the consumable. In some embodiments, the degassing component comprises a cooling component to cool one or more reagents present in the consumable thereby reducing the air present in one or more reagents in the consumable.

[0207] The imaging stations may comprise any of the imaging components described above.Scaling or combining automated high-capacity assay processing units;

[0208] In embodiments, automated assay processing units are configured such that they are capable of being connected to one or more different automated assay processing units.

[0209] In some cases, automated high-capacity assay processing units can be operably connected in parallel. For example, in some cases, the one or more different automated high-capacity assay processing units are adjacent to the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are parallel to the automated high- capacity assay processing unit. In other cases, automated high-capacity assay processing units can be operably connected in series. For example, the one or more different automated high- capacity assay processing units are in series to the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are connected to the automated high-capacity assay processing unit at an end.

[0210] In still other cases, the one or more different automated high-capacity assay processing units are vertically offset from the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are stacked above or below the automated high-capacity assay processing unit. In certain cases, the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.Consumable;[0021 1 ] Embodiments of automated high-capacity assay processing units are configured to accept and operate on a plurality of consumables simultaneously. Embodiments of automated high- capacity assay processing units are configured to accept and operate on consumables that are identical to consumables utilized in embodiments of standard assay processing units or automated medium-capacity assay processing units in addition to consumables that are specifically designed for the high-capacity assay processing unit.METHODS OF DETECTING A TARGET ANALYTE

[0212] The present disclosure provides methods of detecting a target analyte in a sample using the automated assay processing units and consumables disclosed herein. The samples that may be used with the methods will be discussed first followed by the types of analytes followed by the specific types of assay (e.g., digital analysis, immunoassays, nucleic acid analysis, clinical chemistry, etc.) to be used in the methods.I. Samples

[0213] Aspects of the present disclosure automated assay processing units that may be used to process and analyze different analytes or different types of analytes present in a biological sample and methods of use thereof. In some embodiments, the automated assay processing units of the present disclosure comprise fixed or variable primary zones where the sample may be deposited. In order to analyze a sample, more specifically analytes in a sample, the sample is derived from one or more of various sample sources described in this section.Sample Type

[0214] As used herein, "sample", "test sample", or "biological sample" refers to a sample containing or suspected of containing an analyte. For example, International Publication No. WO 2016 / 161400 are incorporated by reference herein and samples of the present disclosure are further described below.

[0215] In some embodiments, a sample of the present disclosure is derived from any suitable source. In other embodiments, the sample comprise a liquid, fluent particulate solid, or fluid suspension of solid particles. In certain embodiments, the sample may be a liquid sample or a liquid extract of a solid sample.

[0216] In some cases, the sample may be processed prior to the analysis described herein. For example, the sample may be separated or purified from its source prior to analysis: however, in certain embodiments, an unprocessed sample containing the analyte may be assayed directly.

[0217] In some embodiments, the source of the analyte molecule may be synthetic (e.g., produced in a laboratory), the environment (e.g., air, soil, etc.), fluid samples, e.g., water supplies, etc.), an animal, e.g., a mammal, a plant, or any combination thereof.

[0218] In some embodiments, a sample as the source of an analyte is a human bodily substance. The human bodily substance may be a liquid sample or a liquid extract of a solid sample. Nonlimiting embodiments of the human bodily substance is bodily fluid, blood, serum, plasma, urine, saliva, sweat, sputum, semen, mucus, lacrimal fluid, tears, demal fluid, lymph fluid, amnioticfluid, interstitial fluid, intestinal fluid, gastrointestinal fluid, lung lavage, spinal fluid, cerebrospinal fluid, feces, nasal mucus, vaginal discharge, tissue, organ, or like. In some embodiments, tissues may include, but are not limited to, skeletal muscle tissue, liver tissue, lung tissue, kidney tissue, myocardial tissue, brain tissue, bone marrow, cervix tissue, skin, etc. In certain cases, the source of the sample may be an organ or tissue, such as a biopsy sample, which may be solubilized by tissue disintegration / cell lysis.

[0219] In certain embodiments, a sample of the present disclosure is whole blood. Samples for hematology are typically whole blood. The whole blood sample consists of red blood cells, white blood cells, and platelets suspended in a protective yellow liquid known as plasma. In some embodiments, samples for immunoassays and clinical chemistry assays are typically serum or plasma. In some embodiments, the whole blood sample is obtained from a subject. In some embodiments, the subject is a living subject, including an animal and a human.

[0220] In certain embodiments, a sample of the present disclosure is venous blood. As used herein, the term “venous blood” refers to a sample of blood taken from a certain vein and checked for specific substances released by nearby organs and tissues. A higher-than-normal amount of a substance can be a sign of disease in the organ or tissue. In some embodiments, venous blood is collected by a venous blood sampling process. For example, in venous blood sampling, a needle is inserted into a vein to collect a sample of blood for testing.

[0221] In certain embodiments, a sample of the present disclosure is capillary blood. As used herein, “capillary blood”, or “capillary sample” refers to a blood sample collected by pricking the skin. Capillary blood is generally obtained by pricking a finger in adults and a heel in infants and small children. Capillaries are tiny blood vessels near the surface of the skin. Capillary plasma typically contains higher concentrations of proteins, calcium and chloride, and lower levels of potassium, sodium, and urea nitrogen compared to venous plasma.

[0222] In certain embodiments, a sample of the present disclosure is plasma. As used herein, the term “plasma” refers to the colorless fluid part of blood, lymph, or milk, in which corpuscles or fat globules are suspended. As such, plasma is the blood's liquid component and is made up of water, proteins, waste products, minerals, clotting factors, immunoglobulins, carbon dioxide and hormones. The method for separating plasma from blood is well known in the art. In exemplary embodiments, plasma is produced when whole blood is collected in tubes that are treated with an anticoagulant. The blood does not clot in the plasma tube, thereby the cells are removed by centrifugation. The supernatant, designated plasma is carefully removed from the cell pellet.

[0223] Tn certain embodiments, a sample of the present disclosure is serum. As used herein, the term “serum” refers to the watery, clear portion of an animal fluid or plant sap. As used herein, the term “blood serum” refers to an amber-colored, protein-rich liquid that separates out whenblood coagulates. In certain embodiments, serum includes, but not limited to, blood serum, serous (or serosal) fluid secreted by the serous glands, and plant sap. The method for separating serum from blood is well known in the art. In exemplary embodiments, the blood serum is collected after whole blood is allowed to clot. The clot is removed by centrifugation, and the resulting supernatant, designated serum, is carefully removed.

[0224] In certain embodiments, a sample of the present disclosure is a cerebrospinal fluid. The term “cerebrospinal fluid (CSF)” refers to a clear fluid that surrounds and protects the brain and spinal cord. The analysis for cerebrospinal fluid may look for proteins, sugar (glucose), and other substances. The method for collecting cerebrospinal fluid is well known in the art. In exemplary embodiments, cerebrospinal fluid is usually obtained through a lumbar puncture (spinal tap). During the procedure, a needle is inserted usually between the 3rd and 4th lumbar vertebrae and the CSF fluid is collected for testing.

[0225] In certain embodiments, a sample of the present disclosure is saliva. As used herein, the term “saliva” refers to watery liquid secreted into the mouth by glands, providing lubrication for chewing and swallowing, and aiding digestion. Saliva consists of 99% water and 1% protein and salts. The method of collecting saliva is well known in the art. In some embodiments, saliva sample can be refrigerated for up to a week before it needs to be added to the stabilizing fluid in the tube.

[0226] In certain embodiments, a sample of the present disclosure is urine. As used herein, the term “urine” refers to a watery, typically yellowish fluid stored in the bladder and discharged through the urethra. Urine is one of the body's chief means of eliminating excess water and salt, and also contains nitrogen compounds such as urea and other waste substances removed from the blood by the kidneys. Collecting a urine sample is well known in the art. In exemplary embodiments, either a "first-catch" or a "mid-stream" sample of urine is collected in a completely sterile container. The first-catch urine sample is the first part of the urine that comes out. The mid-stream urine is for reducing the risk of the sample being contaminated with bacteria from hands, or the skin around the urethra or the tube that carries urine out of the body. In some embodiments, the collected urine sample may be stored in a fridge at 4 °C less than 24 hours in a sealed plastic bag. In certain embodiments, the urine sample is used for infections such as urinary tract infection (UTI), some sexually transmitted infections (STIs) such as chlamydia in men, or kidney damage, such as ACR test.

[0227] In certain embodiments, a sample of the present disclosure is interstitial fluid. As used herein, the expression “interstitial fluid (ISF)”, “lymph”, or “tissue fluid” refers to clear fluid that occupies the space between the cells in the body or fluid found in the spaces around cells. It comes from substances that leak out of blood capillaries. Interstitial fluid helps bring oxygen andnutrients to cells and to remove waste products from them. As new interstitial fluid is made, it replaces older fluid, which drains towards lymph vessels. The method for collecting interstitial fluid is well known in the art. In one embodiment, ISF can typically be collected from skin using suction blisters by applying suction to skin at elevated temperature for up to 1 hr to create blisters filled with ISF.

[0228] In certain embodiments, a sample of the present disclosure is intestinal fluid. Intestinal fluid or gastrointestinal fluid contains, for example electrolytes, bile salts, lipids and lipid digestion products, cholesterol, proteins, enzymes plus other components and may also vary depending upon the anatomical location (stomach vs small intestine vs colon). The method of collecting intestinal fluid samples is well known in the art. In certain embodiments, the intestinal fluid can be collected through a nasojejunal tube and be made into capsules using the freeze-dried powder method.

[0229] In certain embodiments, a sample of the present disclosure is a sample collected from nasal swabs. In certain embodiments, a sample of the present disclosure is a sample collected from throat swabs. In certain embodiments, a sample of the present disclosure is a sample collected from vaginal swabs. Nasal swabs, throat swabs, and vaginal swabs are well known in the art.

[0230] In certain embodiments, a sample includes respiratory specimen. For example, the respiratory specimen includes, but not is limited to, nasal swab, throat swab, sputum, tracheal / bronchial secretion, and bronchial lavage fluid. In some embodiments, respiratory sampling includes upper respiratory materials and lower respiratory secretions. In some cases, the upper respiratory materials comprise nasal swab, throat swab, and the like. In other cases, the lower respiratory secretions comprise sputum, tracheal / bronchial secretion, bronchoalveolar lavage fluid, and the like. In some embodiments, the sputum is collected by well known process in the art. For example, collecting sputum follows the steps of i) taking a very deep breath and holding the air for 5 seconds; ii) slowly breathing out; iii) taking another deep breath and coughing hard until some sputum coming up into mouth; iv) spiting the sputum into a sample container. In some embodiments, tracheal / bronchial secretion is collected by inserting suction catheter as deeply as possible and aspirating secretion, which is well known in the art. In some embodiments, bronchoalveolar lavage fluid is collected by use of bronchoscopy, which is well known in the art.

[0231] In some embodiments, a sample includes any tissue obtained from a subject. In other embodiments, a sample includes any cell obtained from a subject. In still other embodiments, a sample includes an extract of any tissue(s) or cell(s) obtained from a subject. The subject is any living subject including a human. Tn some embodiments, tissues may include, but are not limited to skeletal muscle tissue, liver tissue, heart tissue, lung tissue, pancreas tissue, adipose tissue, stomach tissue, gastrointestinal tract tissue, colon tissue, kidney tissue, myocardial tissue, braintissue, breast tissue, nerve tissue, bone marrow, cervix tissue, skin, etc. In some embodiments, cells may include, but are not limited to skeletal muscle cells, liver cells, heart cells, lung cells, pancreas cells, adipose cells, stomach cells, gastrointestinal tract cells, colon cells, kidney cells, myocardial cells, brain cells, breast cells, nerve cells, bone marrow cells, cervix cells, skin cells, etc. In some cases, the sample is tumor or cancer cells. For example, the sample includes, but is not limited to, brain cancer cells, liver cancer cells, pancreas cancer cells, lung cancer cells, breast cancer cells, kidney cancer cells, metastatic cancer cells, ovarian cancer cells, colorectal cancer cells, bladder cancer cells, thyroid cancer cells, lymphoma cells, cervical cancer cells, gynecologic cancer cells, head and neck cancer cells, mesothelioma cells, myeloma cells, skin cancer cells, prostate cancer cells, uterine cancer cells, vaginal and vulvar cancer cells, and the like. In certain cases, the source of the sample may be an organ or tissue, such as a biopsy sample, which may be solubilized by tissue disintegration / cell lysis. In certain embodiments, a sample may be processed prior to performing immunoassay on the sample. For example, the sample may be concentrated, diluted, purified, amplified, etc.II. Target Analytes

[0232] In some embodiments, one or more analytes in a sample may be measured, detected, or assessed by the automated assay processing units of the present disclosure. The sample may be any sample containing or suspected of containing an analyte. As used herein, "analyte", "target analyte", and "analyte" are used interchangeably and refer to the analyte being measured in the automated assay processing units disclosed herein. Examples of analytes provided herein are for illustrative purposes and are not intended to limit the scope of the present disclosure.

[0233] In some embodiments, but not by way of limitation, the analyte may be a pathogen, a prion protein, a cancer cell, a blood component, or a biomolecule. In some cases, the pathogen is, but not limited to, a vims, a bacterium, a fungus, or a protozoan. In some cases, the prion protein may arise from a sporadic prion disease, a genetic prion disease, or an acquired prion disease. In some cases, the cancer cell may be a cancer cell from a tumor or a circulating tumor cell. In some cases, the blood component may be red blood cells, white blood cells, platelets, or proteins found in the blood. In some cases, a biomolecule may be a metabolite, a macromolecule, a protein, or a chemical compound. Any combination of analytes may be measured by the assays of the methods and systems of the present disclosure.

[0234] In some embodiments, one or more analytes may be a cell, such as, circulating tumor cell. In other embodiments, the analyte is a biological cell (e.g., mammalian, avian, reptilian, other vertebrate, insect, yeast, bacterial, cell, etc.). In other embodiments, the analyte may be an infectious agent, such as a bacterium (e.g., Mycobacterium tuberculosis, Staphylococcus aureus,Shigella dysenteriae, Escherichia coli O157:H7, Campylobacter jejuni, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella 08, and Salmonella enteritidis), virus (e.g., retroviruses (such as HIV), herpesviruses, adenoviruses, lentiviruses, Filoviruses (e.g., West Nile, Ebola, and Zika viruses), hepatitis viruses (e.g., A, B, C, D, and E, including surface antigens, core antigens, (such as, for example, Hepatitis B e-antigen (HBeAg), Hepatitis B surface antigen (HBsAg), Hepatitis B core antigen (HBeAg), phosphorylated Hepatitis B core antigen (P-HBcAg), Hepatitis B core -related antigen (HBcrAg)); HPV, Parvovirus, etc.), a parasite, or fungal spores.

[0235] In exemplary embodiments, one or more analytes are tumor or cancer cells. In some cases, a cancer cell may be directly detected, e.g., a nucleic acid or an antigen specific to the cancer cell is detected. In some cases, the presence of a cancer cell may be detected by a change or mutation in a nucleic acid sequence of the cancer cell, including, but not limited to, a SNP, an insertion, a deletion, a chromosome translocation, or gene amplification. In some cases, a cancer cell may be detected by detecting the presence of tumor or cancer markers associated with the cancer cell. In some cases, a cancer cell may be detected by detecting the expression of receptors associated with a cancer cell. In some cases, a cancer cell may be indirectly detected, e.g., metabolic markers associated with the cancer cell can indicate the presence of the cancer cell.

[0236] For example, types of cancer cells that may be detected by assays of the present disclosure include, but are not limited to, carcinoma cells, leukemia cells, lymphoma cells, myeloma cells, sarcoma cells, central nervous system cancer cells, and mesothelioma cells. Specific types of cancer include, but are not limited to, Bone Cancer (includes Ewing Sarcoma and Osteosarcoma and Malignant Fibrous Histiocytoma), Brain Tumors, Breast Cancer, Cervical cancer, Colorectal Cancer, Endometrial Cancer (Uterine Cancer), Esophageal Cancer, Head and Neck Cancer, Hepatocellular (Liver) Cancer, Hodgkin Lymphoma, Kidney (Renal Cell) Cancer, gynecologic cancer cells, vaginal and vulvar cancer cells, Leukemia, Lung Cancer (Non-Small Cell, Small Cell, Pleuropulmonary Blastoma, Pulmonary Inflammatory Myofibroblastic Tumor, and Tracheobronchial Tumor), Lymphoma, Melanoma, Multiple Myeloma / Plasma Cell Neoplasms, Neuroblastoma, Non-Hodgkin Lymphoma, Ovarian Cancer, Pancreatic Cancer, Prostate Cancer, Skin Cancer, Testicular Cancer, Thyroid Cancer. Markers of cancer include, but are not limited to, ALK gene rearrangements and overexpression, Alpha-fetoprotein (AFP), B-cell immunoglobulin gene rearrangement, BCL2 gene rearrangement, Beta-2-microglobulin (B2M), Beta-human chorionic gonadotropin (Beta-hCG), Bladder Tumor Antigen (BTA), BRCA1 and BRCA2 gene mutations, BCR-ABL fusion gene (Philadelphia chromosome), RAF V600 mutations, C-kit / CD117, CA15-3 / CA27.29, CA19-9, CA-125, CA 27.29, Calcitonin, Carcinoembryonic antigen (CEA), CD19, CD20, CD22, CD25, CD30, CD33, Chromogranin A (CgA), Chromosome 17p deletion, Chromosomes 3, 7, 17, and 9p21, Circulating tumor cells ofepithelial origin (CELLSEARCH), Cytokeratin fragment 21-1, Cyclin DI (CCND1) gene rearrangement or expression, Des-gamma-carboxy prothrombin (DCP), DPD gene mutation, EGFR gene mutation, Estrogen receptor (ER)Zprogesterone receptor (PR), FGFR2 and FGFR3 gene mutations, Fibrin / fibrinogen, FLT3 gene mutations, Gastrin, HE4, HER2 / neu gene amplification or protein overexpression, 5-HIAA, IDH1 and IDH2 gene mutations, Immunoglobulins, 1RF4 gene rearrangement, JAK2 gene mutation, KRAS gene mutation, Lactate dehydrogenase, Microsatellite instability (MSI) and / or mismatch repair deficient (dMMR), MYC gene expression, MYD88 gene mutation, Myeloperoxidase (MPO), Neuron-specific enolase (NSE), NTRK gene fusion, Nuclear matrix protein 22, PCA3 mRNA, PML / RARa fusion gene, Prostatic Acid Phosphatase (PAP), Programmed death ligand 1 (PD-L1), Prostate-specific antigen (PSA), ROS 1 gene rearrangement, Soluble mesothelin-related peptides (SMRP), Somatostatin receptor, T-cell receptor gene rearrangement, Terminal transferase (TdT), Thiopurine S- methyltransferase (TPMT) enzyme activity or TPMT genetic test. Thyroglobulin, UGT1A1*28 variant homozygosity, Urine catecholamines: VMA and HVA, Urokinase plasminogen activator (uPA) and plasminogen activator inhibitor (PALI), FoundationOne CDx (FICDx) genomic test, Guardant360 CDx genomic test, 5-Protein signature (0VA1), 17-Gene signature (Oncotype DX GPS test), 21 -Gene signature (Oncotype DX), 46-Gene signature (Prolaris), 70-Gene signature (Mammaprint).

[0237] Furthermore, types of cancer cells that may be detected by assays of the present disclosure include gastric cancer cells (e.g., HGC-27 cells); non-small cell lung cancer (NSCLC) cells, colorectal cancer cells (e.g., DLD-1 cells), H23 lung adenocarcinoma cells, Ramos cells, T-cell acute lymphoblastic leukemia (T-ALL) cells, CCRF-CEM cells, acute myeloid leukemia (AML) cells (e.g., HL60 cells), small-cell lung cancer (SCLC) cells (e.g., NCI-H69 cells), human glioblastoma cells (e.g., U118-MG cells), prostate cancer cells (e.g., PC-3 cells), HER-2- overexpressing human breast cancer cells (e.g., SK-BR-3 cells), pancreatic cancer cells (e.g., Mia- PaCa-2)).

[0238] In exemplary embodiments, one or more analytes is a virus. In some cases, the vims is directly detected, e.g., a nucleic acid or an antigen specific to the vims is detected. In some cases, the vims is indirectly detected, e.g., detection of anti-vims antibodies produced by a subject can indicate the presence of a vims, or the presence of a vims induces hemagglutination in blood. For example, vimses that may be detected by the assays of the present disclosure include animal, plant, fungal and bacterial vimses. In other embodiments, vimses that may be detected by the assays of the present disclosure include those which impact animals, especially mammals, in particular humans and domestic animals. In still other embodiments, vimses that may be detected by the assays of the present disclosure include, but are not limited to, Papovavimses, e.g. polyoma vimsand SV40: Poxviruses, e.g. vaccinia virus and variola (smallpox); Adenoviruses, e.g., human adenovirus; Herpesviruses, e. g. Human Herpes Simplex types I and II; Parvoviruses, e.g. adeno associated virus (AAV); Reoviruses, e.g., rotavirus and reovirus of humans; Picomaviruses, e.g. poliovirus; Togaviruses, including the alpha viruses (group A), e.g. Sindbis virus and Semliki forest vims (SFV) and the flaviviruses (group B), e.g. dengue vims, yellow fever vims and the St. Louis encephalitis vims; Retrovimses, e. g. lentivimses, HIV I and II, Rous sarcoma vims (RS V), and mouse leukemia viruses; Rhabdovimses, e.g. vesicular stomatitis vims (VSV) and rabies vims; Paramyxoviruses, e.g. mumps vims, measles vims and Sendai vims; Arena viruses, e.g., lassa vims; Bunyavimses, e.g., bunyawere (encephalitis); Coronavimses, e.g. common cold, GI distress vimses, Orthomyxovirus, e.g., influenza; Calicivimses, e.g., Norwak vims, Hepatitis E vims; Filovimses, e.g., ebola vims and Marburg vims; and Astrovimses, e.g. astrovirus, among others. Specific examples of vimses include, but are not limited to, Sin Nombre vims, influenza (especially H5N1 influenza), Herpes Simplex Vims (HSV1 and HSV-2), Coxsackie vims, Human immunodeficiency vims (I and II), Andes vims, Dengue vims, Epstein-Barr vims (mononucleosis), Variola (smallpox) and other pox vimses, West Nile vims, hepatitis vimses (e.g., A, B, C, D, and E, including surface antigens, core antigens, (such as, for example, Hepatitis B e-antigen (HBeAg), Hepatitis B surface antigen (HBsAg), Hepatitis B core antigen (HBcAg), phosphorylated Hepatitis B core antigen (P-HBcAg), Hepatitis B core-related antigen (HBcrAg)), HPV, SARS-CoV-2 (COVID-19), CMV, Parvovims B19, Chlamydia, Gonorrhea, Zika Vims, Chikungunya Vims, Babesia, Malaria, and Usutu vims.

[0239] In some embodiments, one or more analytes may be a bacterium. In some cases, the bacterium is directly detected, e.g., a nucleic acid or an antigen specific to the bacterium is detected. In some cases, the bacterium is indirectly detected, e.g., detection of anti-bacteria antibodies produced by a subject can indicate the presence of bacteria, or the presence of bacterial enzyme activity products can indicate the presence of bacteria. For example, bacteria that may be detected by assays of the present disclosure include, but are not limited to, Achromobacter denitrificans, Achromobacter xylosoxidans, Acinetobacter baumannii, Acinetobacter calcoaceticus, Actinomyces israelii, Aerococcus christensenii, Aeromonas hydrophile, Aeromonas sobria, Aggregatibacter actinomycetemcomitans, Alcaligenes faecalis, Alistipes onderdonkii, Anaerococcus vaginalis, Anaeroglobus geminatus, Arcanobacterium haemolyticum, Arcanobacterium pyogenes, Arthrobacter cumminsii, Atopobium vaginae, Bacillus anthracis, Bacillus cereus, Bacillus coagulans, Bacillus licheniformis, Bacillus megaterium, Bacillus pumilus, Bacillus sphaericus, Bacillus subtilis, Bacteroides dorei, Bacteroides finegoldii, Bacteroides fragilis, Bacteroides nordii, Bacteroides salyersiae, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides vulgatus, Bartonella henselae, Bartonella quintana,Bifidobacterium bifidum, Bifidobacterium breve, Bilophila wadsworthia, Bordetella pertussis, Borrelia burgdorferi, Borrelia recurrentis, Brevibacillus laterosporus, Brucella abortus, Brucella melitensis, Brucella suis, Burkholderia cepacia, Burkholderia mallei, Burkholderia pseudomallei, Campylobacter coli, Campylobacter curvus, Campylobacter jejuni, Campylobacter rectus, Capnocytophaga gingivalis, Capnocytophaga granulosa, Capnocytophaga haemolytica, Capnocytophaga sputigena, Cardiobacterium hominis, Chryseobacterium meningosepticum, Citrobacter amalonaticus, Citrobacter freundii, Citrobacter koseri, Clostridium butyricum, Clostridium difficile, Clostridium histolyticum, Clostridium hylemonae, Clostridium paraputrificum, Clostridium perfringens, Clostridium septicum, Clostridium sporogenes, Clostridium subterminale, Clostridium tertium, Clostridium tetani, Corynebacterium amycolatum, Corynebacterium confusum, Corynebacterium diphtheriae, Corynebacterium glucuronolyticum, Corynebacterium jeikeium, Corynebacterium kroppenstedtii, Corynebacterium macginleyi, Corynebacterium minutissimum. Corynebacterium pseudodiphtheriticum, Corynebacterium pseudotuberculosis, Corynebacterium riegelii, Corynebacterium tuberculostearicum, Corynebacterium ulcerans, Corynebacterium xerosis, Edwardsiella tarda, Eggerthella lenta, Eikenella corrodens, Elizabethkingia meningoseptica, Empedobacter brevis, Enterobacter aerogenes, Enterobacter aerogenes, Enterobacter cloacae, Enterobacter sakazakii, Enterococcus avium, Enterococcus bovis, Enterococcus casseliflavus, Enterococcus cecorum, Enterococcus dispar, Enterococcus durans, Enterococcus faecium, Enterococcus flavescens, Enterococcus gallinarum, Enterococcus gilvus, Enterococcus hirae, Enterococcus italicus, Enterococcus malodoratus, Enterococcus mundtii, Enterococcus pallens, Enterococcus pseudoavium, Enterococcus raffinosus, Enterococcus sanguinicola, Erysipelothrix rhusiopathiae, Escherichia albertii, Escherichia coli, Eubacterium lentum, Eubacterium limosum, Finegoldia magna, Franci sella tularensis, Fusobacterium necrophorum, Fusobacterium nucleatum, Fusobacterium periodonticum, Fusobacterium varium, Gardnerella vaginalis, Gemella morbillorum, Geobacillus stearothermophilus, Granulicatella adiacens, Haemophilus ducreyi, Haemophilus influenzae, Haemophilus parainfluenzae, Hafnia alvei, Halomonas venusta, Helicobacter cinaedi, Helicobacter pylori, Kingella kingae, Klebsiella granulomatis, Klebsiella oxytoca, Klebsiella pneumoniae, Lactobacillus acidophilus, Lactobacillus crispatus, Lactobacillus delbrueckii, Lactobacillus jensenii, Lactococcus garvieae, Legionella pneumophila, Leptospira interrogans, Listeria monocytogenes, Micrococcus luteus, Moraxella catarrhalis, Morganella morganii, Mycoplasma genitalium, Mycoplasma hominis, Neisseria gonorrhoeae, Neisseria meningitidis, Nocardia cyriacigeorgica, Odoribacter splanchnicus, Pantoea agglomerans, Parabacteroides distasonis, Parvimonas micra, Pasteurella multocida, Pediococcus damnosus, Peptoniphilus asaccharolyticus, Peptoniphilus gorbachii, Peptostreptococcus anaerobius, Plesiomonasshigelloides, Porphyromonas asaccharolytica, Porphyromonas gingivalis, Prevotella bivia, Prevotella bivia, Prevotella corporis, Prevotella intermedia, Prevotella melaninogenica, Prevotella nigrescens, Prevotella timonensis, Prevotella veroralis, Propionibacterium acnes, Propionibacterium avidum, Propionibacterium granulosum, Proteus mirabilis, Proteus vulgaris, Providencia rettgeri, Providencia stuartii, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas putida, Rothia dentocariosa, Rothia mucilaginosa, Salmonella enterica, Serratia marcescens, Serratia plymuthica, Shigella boydii, Shigella dysenteriae, Shigella flexneri, Shigella sonnei, Spirillum minus, Staphylococcus aureus, Staphylococcus auricularis, Staphylococcus capitis. Staphylococcus caprae, Staphylococcus carnosus, Staphylococcus cohnii, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus lugdunensis, Staphylococcus pasteuri, Staphylococcus pettenkoferi, Staphylococcus pulvereri, Staphylococcus saccharolyticus, Staphylococcus saprophyticus, Staphylococcus schleiferi, Staphylococcus simulans, Staphylococcus wameri, Staphylococcus xylosus, Stenotrophomonas maltophilia, Streptobacillus moniliformis, Streptococcus agalactiae, Streptococcus anginosus, Streptococcus bovis, Streptococcus canis, Streptococcus constellates, Streptococcus dysgalactiae, Streptococcus equi, Streptococcus gallolyticus, Streptococcus gordonii, Streptococcus infantarius, Streptococcus iniae, Streptococcus intermedins, Streptococcus lutetiensis, Streptococcus mitis, Streptococcus mutans, Streptococcus oralis, Streptococcus pasteuriamis, Streptococcus pneumoniae, Streptococcus porcinus, Streptococcus pyogenes, Streptococcus salivarius, Streptococcus salivarius, Streptococcus sanguinis, Streptococcus sobrinus, Streptococcus suis, Streptococcus vestibularis, Sutterella wadsworthensis, Treponema pallidum, Ureaplasma parvum, Vagococcus fluvialis, Veillonella atypica, Veillonella parvula, Vibrio alginolyticus, Vibrio cholerae, Vibrio fluvialis, Vibrio parahaemolyticus, Vibrio vulnificus, Yersinia enterocolitica, Yersinia pestis, Yersinia pseudotuberculosis.

[0240] In exemplary embodiments, one or more analytes may be a fungus. In some cases, the fungus is directly detected, e.g., a nucleic acid or an antigen specific to the fungus is detected. In some cases, the fungus is indirectly detected, e.g., a cell wall component of a fungus released into the blood can indicate the presence of the fungus. In certain embodiments, fungi that may be detected by assays of the present disclosure include, but are not limited to, fungi from a fungal genera selected from the group consisting of Candida, Aspergillus, Rhyzopus, Cryptococcus, Histoplasma, Pneumocystis, Stachybotrys, Sporothrix, Trichophyton, Microsporum, Blastomyces, Mucoromycotina, Coccidioides, Exserohilum, Cladosporium, Coccoides, Encephalitozoon, Encephalitozoon, Fusarium, Lichtheimia, Mortierella, Malassezia, Prototheca, Pythium, Rhodotorula, Fusarium, Thielaviopsis, Verticillium, Magnaporthe, Sclerotinia, Ustilago, Rhizoctonia, Puccinia, Armillaria, Botrytis, Blumeria, Mycosphaerella, Colletotrichum,Melampsora, Saprolegniasis, Ichthyosporidium, Exophiala, Branchiomycosis, and Penicillium. Specific examples of fungal species that can be detected by the assays of the present disclosure include, but are not limited to, Candida albicans, C. glabrata, C. parapsilosis, C. tropicalis, and C. auris; Cryptococcus neoformans and C. gattii; Coccidioides immitis and C. posadasii; Histoplasma capsulatum; Blastomyces dermatitidis; and Pneumocystis jirovecii.

[0241] In some embodiments, one or more analytes may be a protozoa. In some cases, the protozoan is directly detected, e.g., a nucleic acid or an antigen specific to the protozoan is detected. In other cases, the protozoan is indirectly detected, e.g., a metabolic product of the protozoan can indicate the presence of the protozoan. In some cases, classes of protozoa that may be detected by assays of the present disclosure include, but are not limited to, Plasmodium (malaria), Leishmania (leishmaniasis), Trypanosoma (sleeping sickness and Chagas disease), Cryptosporidium, Giardia, Toxoplasma, Babesia, Balantidium and Entamoeba. Specific examples of protozoa that can be detected by the assays of the present disclosure include, but are not limited to, Plasmodium falciparum, Plasmodium ovale, Plasmodium malariae, Plasmodium vivax, Leishmania donovani, Trypanosoma brucei, Trypanosoma cruzi, Toxoplasma gondii and Babesia microti.

[0242] In exemplary embodiments, one or more analytes may be a prion protein. In some cases, the prion is directly detected, e.g., a nucleic acid or an antigen specific to the prion is detected. In other cases, the presence of prions or potential for prion formation is detected by identifying a mutation in a nucleic acid sequence. In some cases, the presence of structures formed by prions can indicate the presence of prions. In some cases, the prion is indirectly detected, e.g., biochemical changes induced by prion formation can indicate the presence of prions. In some cases, prions are amplified prior to detection using methods such as protein misfolding cyclic amplification (PMCA) or real-time quaking-induced conversion (RT-QUIC). Exemplary prion proteins include, but are not limited to, Scrapie (Sheep and goats), transmissible mink encephalopathy (TME), chronic wasting disease (CWD) in mule deer and elk, bovine spongiform encephalopathy (BSE) cattle, feline spongiform encephalopathy (FSE) in cats, exotic ungulate encephalopathy (EUE), Kuru in humans, Creutzfeldt-Jakob disease (CJD) in humans, Fatal familial insomnia (FFI) in humans and Gerstmann-Strassler-Scheinker syndrome (GSS) in humans.

[0243] In some embodiments, one or more analytes measured by the assays of the automated assay processing units and methods of the present disclosure may be a blood component. Examples of blood components that may be detected by assays of the present disclosure include, but are not limited to, red blood cells, hemoglobin, white blood cells (including neutrophils, lymphocytes, monocytes, eosinophils, and basophils), platelets, reticulocytes, and nucleated redblood cells. Various measurements of different blood components may be performed, including, but not limited to, cell count, cell size, cell complexity, granularity, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration.

[0244] In some embodiments, one or more analytes may be a biomolecule. Non-limiting examples of biomolecules include macromolecules such as, for example, proteins, lipids, and carbohydrates. In certain instances, the analyte may be hormones, antibodies, growth factors, cytokines, electrolytes (e.g., sodium, potassium, and chloride), enzymes (e.g., alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase, and amylase), receptors (e.g., neural, hormonal, nutrient, and cell surface receptors) or their ligands, cancer markers (e.g., PSA, TNF-alpha), markers of myocardial infarction (e.g., troponin, creatine kinase, B-type natriuretic peptide (also known as brain natriuretic peptide; BNP), N-terminal prohormone of brain natriuretic peptide (NT-proBNP) and the like), toxins, drugs (e.g., therapeutic drugs, drugs of addiction), metabolic agents (e.g., including vitamins and minerals), metabolic products (e.g., glucose, urea nitrogen triglycerides, uric acid), nutrients, and the like. Non-limiting embodiments of protein analytes include peptides, polypeptides, protein fragments, protein complexes, fusion proteins, recombinant proteins, phosphoproteins, glycoproteins, lipoproteins, and the like. In certain embodiments, the analyte may be a post-translationally modified protein (e.g., phosphorylated, methylated, glycosylated protein). In certain embodiments, the analyte is a nucleic acid. In certain embodiments, the analyte is a protein or a small molecule.

[0245] A non-limiting list of analytes that may be analyzed by the automated assay processing units presented herein include anti-Mullerian Hormone (AMH), autoantibody to CD25, Chemokine (C-X-C) motif ligand 13 (CXCL13), Dickkopf-3 (Dkk-3), IL-12p40, Interleukein 8 (IL-8), pl4 Endocan Fragment, SARS-CoV-2 IgA Antibody, SARS-CoV-2 IgG Antibody, SARS- CoV-2 IgM Antibody, Secretory Gelsolin (pGSN), Secretogranin II, ACE2, Albumin, Albuminuria, Alpha-Amylase, Apo H, B-2 Microglobulin, Brain natriuretic peptide (BNP) and derivatives thereof (e.g., preproBNP, proBNP, NT-proBNP, BNP (1-32)), CA 24-2, Carcinoembryonic Antigen (CEA), Cardiac myosin binding protein C, Ceruloplasmin, Cyclosporine, C-peptide, C-Reactive Protein (CRP), Dipeptidyl peptidase 4 (DPP -4), Digoxin, Fibrinogen Alpha Chain (FGA), Homocysteine, Interleukin 18 (IL-18), Interleukin 6 (IL-6), Lactate Dehydrogenase (LD), Liver Fatty Acid Binding Protein (L-FABP), Lipase, Microalbuminuria, Neutrophil Gelatinase-Associated Lipocalin (NG AL), Osteopontin, Periostin, Peroxisome Proliferator- Activated Receptor Gamma Coactivator-1 Alpha (PGC-la), Proapoptotic Kinase R (PKR) and its phosphorylated PKR (pPKR), Procalcitonin (PCT), Pepsinogen I, Pepsinogen II, Pro-SFTPB, PTH (Parathyroid Hormone), soluble Interleukin 2 (sIL- 2), Sex Hormone-Binding Globulin (SHBG), Thioredoxin, TSH (Thyroid Stimulating Hormone),Vitamin D-Binding Protein, Alpha-synuclein, BARF1 (BamHl-A Reading Frame 1), Kidney Injury Molecule-1 (KIM-1), Laminin gamma (e.g., laminin gamma subunit 1, laminin gamma subunit 2, laminin gamma subunit 3), LMP1 (Latent Membrane Protein 1), Neurofilament light chain (NF-L), Tau protein, Tau, UCH-L1 (Ubiquitin C-Terminal Hydrolase-Ll), Alkaline Phosphatase, Amylase, Aspartate Aminotransferase (AST), Calcium, Cholesterol, Creatine Kinase (CK), Carbon Dioxide (C02), Creatinine, Direct Low-Density Lipoprotein (Direct LDL), Gamma-Glutamyl Transferase (GGT), High-Density Lipoprotein (HDL), Iron, Low-Density Lipoprotein (LDL), Magnesium, Potassium (K), Sodium (Na), Triglycerides, Uric Acid, Akt (Protein Kinase B), Amphiregulin, ANXA7 (Annexin A7), Androgen Receptor (AR), v-Raf Murine Sarcoma Viral Oncogene Homolog B (BRAF), Cyclin-Dependent Kinase Inhibitor IB (CDKN1B), MYC Proto-Oncogene (cMYC), Catenin Beta-1 (CTNNB1), Epidermal Growth Factor Receptor (EGFR), Ephrin Type-B Receptor 2 (EPHB2), Estrogen Receptor 1 (ESRI), Estrogen Receptor 2 (ESR2), Ferritin, Folate, Forkhead Box 03 (F0X03A), Mechanistic Target of Rapamycin Complex 1 (FRAP1), Fibroblast Growth Factor Receptor Substrate 2 (FRS2), GRB2-Associated Binding Protein 2 (Gab2), Glial Fibrillary Acidic Protein (GFAP), Growth Factor Receptor-Bound Protein 2 (Grb2), Growth Regulation By Estrogen In Breast Cancer 1 (GREB1), Hepatitis B e-antigen (HBeAg), Hepatitis B surface antigen (HBsAg), Hepatitis B core antigen (HBeAg), phosphorylated Hepatitis B core antigen (P-HBcAg), Hepatitis B core-related antigen (HBcrAg), Human Epidermal Growth Factor Receptor 2 (HER2), Human Epidermal Growth Factor Receptor 3 (HER3), Human Epidermal Growth Factor Receptor 4 (HER4), Insulin- Like Growth Factor 1 Receptor (IGF-IR), IL6R (Interleukin-6 Receptor), Kruppel Like Factor 6 (KLF6), Kirsten Rat Sarcoma Viral Oncogene Homolog (KRAS), Leucine Zipper Tumor Suppressor 1 (LZTS1), Mitogen- Activated Protein Kinase Kinase 1 (MAP2K1), Mitogen- Activated Protein Kinase (MEK), Mitogen-Inducible Gene 6 Protein (MIG-6), Marker of Proliferation Ki-67 (MKI67), Mechanistic Target of Rapamycin (mTOR), Mucin 4, Cell Surface Associated (MUC4), Neural Precursor Cell Expressed Developmentally Down-Regulated Protein 4-1 (NEDD4-1), NK3 Homeobox 1 (NKX3-1), Neuregulin 1 (NRG1), Parkin, Parvovirus B 19, 3- Phosphoinositide-Dependent Protein Kinase-1 (PDK-1), Progesterone Receptor (PGR), PH Domain and Leucine -Rich Repeat Protein Phosphatase (PHLPP), Phosphatidylinositol-4,5- Bisphosphate 3-Kinase Catalytic Subunit Alpha (PIK3CA), Paired-Like Homeodomain Transcription Factor 2 (PITX2), Protein Phosphatase 1 Regulatory Inhibitor Subunit IB (PPP1R1B), PRDX6 (Peroxiredoxin-6), Phosphatase and Tensin Homology (PTEN), Phosphatase and Tensin Homolog 1 (PTEN 1), PXN (Paxillin), Ribosomal Protein S6 Kinase (S6K), Src Homology 2 Domain-Containing Inositol Phosphatase (SHIP), Sirolimus, Src Proto-Oncogene Tyrosine-Protein Kinase Src (Src), Tacrolimus, Thyroxine (Total or Free T4), Triiodothyronine(Total or Free T3), Thyroglobulin, DNA Topoisomerase II (TOPO II), TRAb, Tuberous Sclerosis 1 (TSC1), Tuberous Sclerosis 2 (TSC2), Tumor necrosis factor-alpha receptors. Amyloid BetaProtein 42 (A042), Creatine Kinase-MB (CK-MB), Anti-Cyclic Citrullinated Peptide (Anti-CCP), Anti-Thyroglobulin Antibody (Anti-Tg), Anti-Thyroid Peroxidase Antibody (Anti-TPO), Antistreptolysin O (ASO), Complement Component 3 (C3), Complement Component 4 (C4), D- Dimer, Rheumatoid Factor (RF), DJ-1, Leucine-rich repeat kinase 2, Mutated ATP13A2, Phenobarbital, Phenytoin, Prion protein, PTEN induced putative kinase 1, Alpha-Fetoprotein (AFP), CA 125 (MUC16), CA 15-3, CA 19-9, Cyclin I (CCNI), Cytomegalovirus (CMV), CYFRA21-1, Fibroblast Growth Factor 19 (FGF19), Gentamicin, Human Epididymis Protein 4 (HE-4), Neuron-Specific Enolase (NSE), Perinuclear Anti-Neutrophil Cytoplasmic Antibody (p- ANCA), Protein Induced By Vitamin K Absence (PIVKA), Protein Induced By Vitamin K Absence-II (PIVKA-II), Pro-Surfactant B (Pro-SFTPB), Prostate Specific Antigen (PSA), Rubella, Squamous Cell Carcinoma Antigen (SCC), Toxo IgG, Toxo IgM, Beta Human Chorionic Gonadotropin (Beta-hcG), Botulinum toxins, Clostridium difficile toxins A and B, Dehydroepiandrosterone Sulfate (DHEA-S), Diphtheria toxin, E. coli enterotoxins (heat-labile exotoxin, heat-stable enterotoxin), Fetuin-A, Follicle-Stimulating Hormone (FSH), Glycosylated hemoglobin (HbAlc), Hemoglobin Ale, Interleukin 1 alpha (ILla), Influenza HA antigen, Luteinizing Hormone (LH), Methotrexate, Myeloperoxidase (MPO), Neurofibromin 1 (NF-1), Plasma C-peptide, Placental Growth Factor (P1GF), Pro-GFP, Prolactin, S 1000, Soluble Fms-Like Tyrosine Kinase-1 (sFlt-1), Testosterone, Tetanus toxin, Thymosin 1315, Alanine Aminotransferase (ALT), Bile Acids-Total, Bilirubin, Bilirubin-Direct, Bilirubin-Total, Calprotectin, Deoxyuridine Triphosphatase (DUTPase), Lactic Acid, Lactoferrin, Shiga toxin, Shiga-like toxin I, Shiga-like toxin II, Theophylline, Total Protein, Urea Nitrogen, Valproic acid, Vitamin B12, Voltage-Dependent Anion Channel 1 (VDAC1), Wilm's Tumor-1 protein, Amphetamines, Methamphetamines, Barbiturates, Benzodiazepines, Benzodiazepines (Seram), Cannabinoids, Ecstasy, Ethanol, Opiates, or Phencyclidine (PCP) or any combination thereof.

[0246] Examples of nucleic acid aptamers include: drags of abuse (e.g. cocaine), protein biomarkers (including, but not limited to, Nucleolin, nuclear factor-kB essential modulator (NEMO), CD-30, protein tyrosine kinase 7 (PTK7), vascular endothelial growth factor (VEGF), MUC1 glycoform, immunoglobulin p Heavy Chains (IGHM), Immunoglobulin E, av03 integrin, a-thrombin, NF-KB, E2F transcription factor, Plasminogen activator inhibitor, Tenascin C, CXCL12 / SDF-l , prostate specific membrane antigen (PSMA), gastric cancer cells, HGC-27 ; cells (including, but not limited to, non-small cell lung cancer (NSCLC), colorectal cancer cells, (DLD- 1), H23 lung adenocarcinoma cells, Ramos cells, T-cell acute lymphoblastic leukemia (T-ALL) cells, CCRF-CEM, acute myeloid leukemia (AML) cells (HL60), small-cell lung cancer (SCLC)cells, NCIH69, human glioblastoma cells, U118-MG, PC-3 cells, HER-2-overexpressing human breast cancer cells (SK-BR-3), pancreatic cancer cell line (Mia-PaCa-2)); and infectious agents (including, but not limited to, Mycobacterium tuberculosis, Staphylococcus aureus, Shigella dysenteriae, Escherichia coli O157:H7, Campylobacter jejuni, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella 08, or Salmonella enteritidis).

[0247] Exemplary targets of protein or peptide aptamers that may be measured in a sample obtained from a patient or subject in need using the subject automated assay processing units include, but are not limited to: HBV core capsid protein, CDK2, E2F transcription factor, Thymidylate synthase, Ras, EB1, and Receptor for Advanced Glycated End products (RAGE). Aptamers, and use and methods of production thereof are reviewed in e.g., Shum et al., J Cancer Ther. 2013 4:872; Zhang et al., Curr Med Chem. 201 1;18:4185; Zhu et al., Chem Commun (Camb). 2012 48:10472; Crawford et al., Brief Funct Genomic Proteomic. 2003 2:72; Reverdatto et al., PLoS One. 2013 8:e65180.

[0248] In certain cases, a biological sample (e.g., human blood sample) that contains or is suspected of containing a target nucleic acid may undergo preparation / processing prior to detection by a primary analysis unit of a system of the present disclosure. In some embodiments, the preparation / processing may include the following steps: i) isolation of total nucleic acid that contains a target nucleic acid from the sample, ii) optionally, enrichment of the target nucleic acid, iii) amplification of the target nucleic acid, and iv) processing of the amplified target nucleic acid. Each step can be performed manually, automatically, or by a combination thereof.

[0249] In certain embodiments, the analyte is not amplified (i.e., the copy number of the analyte is not increased) prior to the measurement of the analyte. For example, in cases where the analyte is DNA or RNA, the analyte is not replicated to increase copy numbers of the analyte.Ill, Types of assays and analysisDigital (individual) analysis

[0250] The automated assay processing units and methods disclosed herein are capable of performing digital analysis of the analytes of the present disclosure. The placement of single microparticles bound to the analyte molecules into wells allows for a digital readout. For example, for a low number of positive wells (<~70% positive) Poisson statistics can be used to quantitate the analyte concentration in a digital format. A digital signal may be used for lower analyte concentrations. As used herein, a “positive well” refers to a well that has a signal related to presence of a microparticle bound to the analyte molecule, which signal is above a threshold value. As used herein, a “negative well” refers to a well that may not have a signal related to presence ofa microparticle bound to the analyte molecule. In certain embodiments, the signal from a negative well may be at a background level, i.e., below a threshold value.

[0251] Because every single analyte, as an end-point entity, can be detected in the context of digital detection, the components and methods associated with digital detection can significantly increase detection sensitivity for sample analysis compared to systems using analog optical detection. As such, using digital detection varying levels of analyte can be differentiated and quantified earlier, decreasing the time to process the sample for detection. Additionally, or alternatively, detection can be performed using a smaller sample volume, less reagent material, less conjugate material, fewer microparticles, or any combination of these, which can reduce costs to perform each assay. As such, and as described herein, sample preparation time can be improved due at least in part to less sample manipulation involved (e.g., faster washing times) and / or improved kinetics of reactions achieved using a lower sample volume, less reagent or conjugate material, ability to use different sample types where the concentration of the targeted analyte may be to low to detect using conventional technology and / or fewer microparticles to obtain an analyte concentration suitable for detection. Assays using less sample volume and / or reagent material can be performed using smaller equipment, which can reduce the footprint of the laboratory system for performing the assays as discussed further herein. In addition, or as a further alternative, increased detection sensitivity can provide additional benefits when used with multiplexing

[0252] Digital detection can provide increased sensitivity due at least in part to a reduction of noise during detection relative to the signal being measured, for example, producing a higher signal-to-noise ratio. Such improved signal-to-noise ratios are possible by coupling the analyte, e.g., a particular nucleic acid, protein, or nucleic acid associated with a protein or microparticles, to an independently detectable end-point entity. For example, but not limitation, amplified nucleic acids or proteins can be immobilized to microparticles and labeled with detectable conjugates, where the conjugate is a detectable end-point entity in that it can emit an independently detectable signal, either directly or via the conversion of a substrate.

[0253] Additionally, or alternatively, and in accordance with another aspect of the disclosed subject matter, the detection operation employs a digital microwell detection process. Additionally, or alternatively, a support medium, such as, but not limited to, microparticles or other labels, can be mixed with the sample in order to perform the digital detection process after amplification. In certain embodiments, reagents including antibodies and coated microparticles can be combined.Immunoassays

[0254] The target analyte, and / or peptides of fragments thereof, may be analyzed using antianalyte antibodies in an immunoassay. The presence or amount can be determined using antibodies and detecting specific binding to the analyte. For example, the antibody, or antibody fragment thereof, may specifically bind to the target analyte.

[0255] The presence or amount of analyte present in a sample may be readily determined using an immunoassay, such as sandwich immunoassay (e.g., monoclonal-monoclonal sandwich immunoassays, monoclonal-polyclonal sandwich immunoassays, including radioisotope detection (radioimmunoassay (RIA)) and enzyme detection (enzyme immunoassay (EIA) or enzyme-linked immunosorbent assay (ELISA) (e.g., Quantikine ELISA assays, R&D Systems, Minneapolis, MN)). An example of a point-of-care automated assay processing unit that can be used is i-STAT® (Abbott, Laboratories, Abbott Park, IL). Other methods that can be used include a chemiluminescent microparticle immunoassay, in particular one employing the ARCHITECT® automated analyzer (Abbott Laboratories, Abbott Park, IL), as an example. Other methods include, for example, mass spectrometry, and immunohistochemistry (e.g., with sections from tissue biopsies), using anti-analyte antibodies (monoclonal, polyclonal, chimeric, humanized, human, etc.) or antibody fragments thereof against analyte. Other methods of detection include those described in, for example, U.S. Patent Nos. 6,143,576; 6,113,855; 6,019,944; 5,985,579; 5,947,124; 5,939,272; 5,922,615; 5,885,527; 5,851,776; 5,824,799; 5,679,526; 5,525,524; and 5,480,792, each of which is hereby incorporated by reference in its entirety. Specific immunological binding of the antibody to the analyte can be detected via direct labels, such as fluorescent or luminescent tags, metals and radionuclides attached to the antibody or via indirect labels, such as alkaline phosphatase or horseradish peroxidase.

[0256] The use of immobilized antibodies or antibody fragments thereof may be incorporated into the immunoassay. The antibodies may be immobilized onto a variety of supports, such as magnetic or chromatographic matrix particles, the surface of an assay plate (such as microtiter wells), pieces of a solid substrate material, and the like.

[0257] A homogeneous format may be used. For example, after the sample is obtained from a subject, a mixture is prepared. The mixture contains the sample being assessed for analyte, a first specific binding partner, and a second specific binding partner. The order in which the sample, the first specific binding partner, and the second specific binding partner are added to form the mixture is not critical. The sample is simultaneously contacted with the first specific binding partner and the second specific binding partner. In some embodiments, the first specific binding partner and any analyte contained in the sample may form a first specific binding partner-analyte- antigen complex and the second specific binding partner may form a first specific binding partneranalyte of interest-second specific binding partner complex. In some embodiments, the secondspecific binding partner and any analyte contained in the sample may form a second specific binding partner-analyte-antigen complex and the first specific binding partner may form a first specific binding partner-analyte of interest-second specific binding partner complex.

[0258] A heterogeneous format may be used. For example, after the sample is obtained from a subject, a first mixture is prepared. The mixture contains the sample being assessed for analyte and a first specific binding partner, wherein the first specific binding partner and any analyte contained in the sample form a first specific binding partner-analyte -antigen complex. The order in which the sample and the first specific binding partner are added to form the mixture is not critical.

[0259] The first specific binding partner may be immobilized on a solid phase. The solid phase used in the immunoassay (for the first specific binding partner and, optionally, the second specific binding partner) can be any solid phase known in the art, such as, but not limited to, microparticle, a bead, a test tube, a microtiter plate, a cuvette, a membrane, a scaffolding molecule, a film, a filter paper, a disc, and a chip. Microparticle may be ferromagnetic, ferrimagnetic, paramagnetic, superparamagnetic or ferrofluidic. Exemplary ferromagnetic materials include Fe, Co, Ni, Gd, Dy, CrO2, MnAs, MnBi, EuO, and NiO / Fe. Examples of ferrimagnetic materials include NiFe2O4, CoFe2O4, Fe3O4 (or FeO.Fe2O3). Microparticles can have a solid core portion that is susceptible to a magnetic field and is surrounded by one or more non-magnetic layers. Alternately, the portion susceptible to a magnetic field can be a layer around a non-magnetic core. The solid support on which the first specific binding member is immobilized may be stored in dry form or in a liquid. The microparticles may be subjected to a magnetic field prior to or after contacting with the sample with a microparticle on which the first specific binding member is immobilized.

[0260] After the mixture containing the first specific binding partner-analyte antigen complex is formed, any unbound analyte is removed from the complex using any technique known in the art. For example, the unbound analyte can be removed by washing. Desirably, however, the first specific binding partner is present in excess of any analyte present in the sample, such that all analyte that is present in the sample is bound by the first specific binding partner.

[0261] After any unbound analyte is removed, a second specific binding partner is added to the mixture to form a first specific binding partner-analyte of interest-second specific binding partner complex.Sandwich assay

[0262] A sandwich immunoassay measures the amount of antigen between two layers of antibodies (i.e., at least one capture antibody) and a detection antibody (i.e., at least one detection antibody). The capture antibody and the detection antibody bind to different epitopes on theantigen, e.g., analyte of interest. Desirably, binding of the capture antibody to an epitope does not interfere with binding of the detection antibody to an epitope. Either monoclonal or polyclonal antibodies may be used as the capture and detection antibodies in the sandwich immunoassay.

[0263] Generally, at least two antibodies are employed to separate and quantify analyte in a sample. More specifically, the at least two antibodies bind to certain epitopes of analyte forming an immune complex which is referred to as a "sandwich". One or more antibodies can be used to capture the analyte in the sample (these antibodies are frequently referred to as a "capture" antibody or "capture" antibodies) and one or more antibodies is used to bind a detectable (namely, quantifiable) label to the sandwich (these antibodies are frequently referred to as the "detection" antibody or "detection" antibodies). In a sandwich assay, the binding of an antibody to its epitope desirably is not diminished by the binding of any other antibody in the assay to its respective epitope. Antibodies are selected so that the one or more first antibodies brought into contact with a sample suspected of containing analyte do not bind to all or part of an epitope recognized by the second or subsequent antibodies, thereby interfering with the ability of the one or more second detection antibodies to bind to the analyte.

[0264] The antibodies may be used as a first antibody in said immunoassay. The antibody immunospecifically binds to epitopes on analyte. In addition to the antibodies of the present disclosure, said immunoassay may comprise a second antibody that immunospecifically binds to epitopes that are not recognized or bound by the first antibody.

[0265] A sample suspected of containing analyte can be contacted with at least one first capture antibody (or antibodies) and at least one second detection antibodies either simultaneously or sequentially. In the sandwich assay format, a sample suspected of containing analyte is first brought into contact with the at least one first capture antibody that specifically binds to a particular epitope under conditions which allow the formation of a first antibody-analyte antigen complex. If more than one capture antibody is used, a first multiple capture antibody-analyte antigen complex is formed. In a sandwich assay, the antibodies, preferably, the at least one capture antibody, are used in molar excess amounts of the maximum amount of analyte expected in the sample. For example, from about 5 pg / mL to about 1 mg / mL of antibody per ml of microparticle coating buffer may be used.

[0266] Provided herein are methods for measuring or detecting a target analyte present in a biological sample. In some embodiments, the target analyte is a protein. In such methods, a consumable comprising wells is loaded into the automated assay processing unit. Following the loading of the consumable, a number of fluids may be dispensed into various regions prior to further processing the sample in an assay. In some embodiments, a sample comprising a target analyte is dispensed into a first region of the consumable. In some embodiments, a particle solutionis dispensed into the first region of the consumable. The particle solution may comprise a single type of particle or multiple types of particles. In some embodiments, the particle solution comprises microparticles. In some embodiments, the particle solution comprises microparticles and assisting microparticles. In some embodiments, a conjugate solution comprising a second specific binding partner that is detectably labeled, e.g., a detection antibody, is dispensed into a second region of the consumable. In some embodiments, a hydrophilic liquid is dispensed into a third region of the consumable comprising wells. In some embodiments, the hydrophilic liquid is a substrate solution. Additionally, wash buffer may be dispensed into at least a fourth region and a fifth region of the consumable. The wash buffer may be dispensed into more than the fourth and the fifth region, such as a sixth, a seventh, an eighth, a nineth, a tenth, an eleventh, a twelfth, a thirteenth, a fourteenth, a fifteenth region, or more than a fifteenth region. Optionally, a lysis buffer may be dispensed into the first region.

[0267] A number of fluids are dispensed into various regions of the consumable. The fluids may be dispensed in any order deemed useful. For instance, the sample, the particle solution, the conjugate solution, the hydrophilic liquid, the wash buffer, and the lysis buffer may be dispensed at the same time or at different times. In some embodiments, the sample is dispensed from a sample dispenser comprising a sample pump and a sample nozzle. In some embodiments, the conjugate solution, the wash buffer, and the lysis buffer may be dispensed from one or more reagent dispensers. The conjugate solution, the wash buffer, and the lysis buffer may be dispensed from the same reagent dispenser or different reagent dispensers. In some embodiments, the hydrophilic liquid is dispensed from a hydrophilic liquid dispenser.

[0268] Following sample and particle solution dispensing, the sample may be incubated with the microparticles or microparticles and assisting microparticles for a time sufficient to allow binding of the first specific binding partner to an analyte present in the sample thereby producing microparticles bound to the target analyte where the microparticles are attached to the first specific binding partner and the first specific binding partner is bound to the target analyte. During the incubation, the consumable may be heated to a desired temperature using a heating element. In some embodiments, the desired temperature is 30-45 C. The sample may be mixed with the microparticles or microparticles and assisting microparticles using a mixing component. In some embodiments, mixing comprises engaging the engagement arm of the mixing component with at least a portion of the consumable and vibrating or agitating the consumable. In some embodiments, the mixing comprises contacting the first region with a ballistic mixing device of a mixing component and agitating the first region.

[0269] Optionally, the microparticles bound to the target analyte or microparticles bound to the target analyte and assisting particles may be subjected to a magnetic field to move themicroparticles bound to the target analyte to a fourth region of the consumable comprising wash buffer. In some embodiments, the subjecting the microparticles bound to the target analyte to a magnetic field comprises positioning the first magnet under the first region, un-retracting the first magnet, moving the first magnet or the consumable positioning the first magnet under the fourth region, and retracting the first magnet. Once in the fourth region, the microparticles or the microparticles and assisting microparticles may be mixed with the wash buffer. In some embodiments, the mixing comprises positioning the second magnet over the fourth region, unretracting the second magnet, and retracting the second magnet.

[0270] Next, the microparticles bound to the target analyte or microparticles bound to the target analyte and assisting particles may be subjected to a magnetic field to move the microparticles bound to the target analyte or microparticles bound to the target analyte and assisting particles to a second region containing a conjugate solution comprising a second specific binding partner (e.g., a detection antibody). The second specific binding partner may be detectably labeled. The label may be any label that can be optically detected. For example, the label may be a fluorescent label or a label that reacts with a substrate solution to produce a detectable label. The microparticles bound to the target analyte may be incubated for a period of time sufficient for the second specific binding partner to bind the analyte bound to the first binding member thereby producing microparticles bound to detectably labeled target analyte. During the incubation the consumable may be heated to a desired temperature using a heating element. In some embodiments, the desired temperature is 30-45 C. The sample may be mixed with the microparticles or microparticles and assisting microparticles using a mixing component. In some embodiments, the mixing comprises positioning the second magnet over the second region, un-retracting the second magnet, and retracting the second magnet. Optionally, the microparticles bound to the detectably labeled target analyte or microparticles bound to the detectably labeled target analyte and assisting particles may be subjected to a magnetic field to move the microparticles to fifth containing wash buffer and mixed.

[0271] Next, the microparticles bound to the detectably labeled target analyte or microparticles bound to the detectably labeled target analyte and assisting particles may be subjected to a magnetic field to move the microparticles to a third region of the consumable comprising wells. The microparticles bound to the detectably labeled target analyte may be allowed to settle into the array of wells in the third region. The microparticles may settle using gravitational force or by applying electric or magnetic force. Next, the wells containing the microparticles bound to the detectably labeled target analyte are sealed using a hydrophobic liquid (e.g., oil). In some embodiments, the sealing comprises dispensing from a hydrophobic liquid dispenser the hydrophobic liquid into the third region thereby flowing the hydrophobic liquid over the wells anddisplacing the hydrophilic liquid and aspirating the hydrophilic liquid using an aspiration device. In some embodiments, the dispensing of the hydrophobic liquid and the aspirating the hydrophilic liquid occurs simultaneously. Lastly, the wells are imaged thereby detecting the detectable signal produced by the hydrophilic liquid reacting with the second specific binding partner. In some embodiments, the imaging of the wells comprises i) immobilizing the consumable, ii) adjusting a focus of an imaging component to a first focal plane, iii) capturing a first scatter image, iv) capturing a first fluorescent image, and v) repeating steps ii-iv) one or more times to generate a stack of scatter images and fluorescent images spanning a plurality of focal planes. In some embodiments, step iii) comprises enabling one or more scatter LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more scatter LEDs. In some embodiments, step iv) comprises enabling one or more fluorescent LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more fluorescent LEDs. In some embodiments, step i) comprises contacting an alignment datum with the consumable thereby activating a raising element and a clamping component, wherein: the raising element is positioned below the consumable and the clamping component is positioned above the consumable, and the raising element provides an upward force on the consumable and the clamping component provides a downward force on the consumable.

[0272] In some instances, it may be desirable to analyze multiple analytes at once. In these embodiments, the microparticles comprise a plurality of first microparticles and a plurality of second microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first epitope and the first specific binding partner of the plurality of second microparticles is specific to a second epitope, and the plurality of first microparticles comprise a first fluorophore and the plurality of second microparticles comprise a second fluorophore. In some embodiments, the microparticles comprise a plurality of first microparticles, a plurality of second microparticles, and a plurality of third microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first epitope, the first specific binding partner of the plurality of second microparticles is specific to a second epitope, the first specific binding partner of the plurality of third microparticles is specific to a third epitope and the plurality of first microparticles comprise a first fluorophore, the plurality of second microparticles comprise a second fluorophore, and the plurality of third microparticles comprise a third fluorophore. In some embodiments, the first fluorophore, the second fluorophore, and the third fluorophore are the same fluorophore. In some embodiments, the first fluorophore, the second fluorophore, and the third fluorophore are different fluorophores. In some embodiments, the second fluorophore and the third fluorophore emits at the same wavelength but are excited by a different wavelength of light. When two or more of the fluorophores are the same fluorophore, the fluorophores may be presentin different concentrations such that the concentration of the first fluorophore is a lower or higher concentration than that of the second fluorophore. In some embodiments, the first epitope, the second epitope, and the third epitope are the same epitope. In some embodiments, the first epitope, the second epitope, and the third epitope are different epitopes.

[0273] When multiple analyte detection is desired, the target analyte may be a first target analyte and a second target analyte, and the first target analyte is different from the second target analyte. In some embodiments, the target analyte is a first target analyte, a second target analyte, and a third target analyte, and the first, second, and third target analyte are different. In some embodiments, the second target analyte is a control analyte. By “control analyte” it is meant that the analyte is used as a control to determine if the assay steps were performed without error and the assay reagents performed as expected. The control analyte may be an analyte that is added to the sample in a known concentration. In an alternative embodiment, the control analyte may be endogenously present in the sample. In some embodiments, the control analyte is biotin. When the control analyte is biotin, the first specific binding partner of the plurality of second microparticles is antibody or fragment thereof that binds to biotin. In some embodiments, the control analyte is biotin. When the control analyte is biotin, the first specific binding partner of the plurality of second microparticles is streptavidin.

[0274] In some embodiments, the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles and / or the plurality of third microparticles. In some embodiments, the plurality of first microparticles are coated with the first fluorophore, the plurality of second microparticles are coated with the second fluorophore, and the plurality of third microparticles re coated with the third fluorophore. In some embodiments, the plurality of first microparticles are embedded with the first fluorophore, the plurality of second microparticles are embedded with the second fluorophore, and the plurality of third microparticles are embedded with the third fluorophore. In some embodiments, capturing the first fluorescent image of step iv) further comprises capturing a second fluorescent and capturing a third fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, and the third fluorescent image detects the plurality of second microparticles. In some embodiments, capturing the first fluorescent image of step iv) further comprises capturing a second fluorescent, capturing a third fluorescent image, and capturing fourth fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, the third fluorescent image detects the plurality of second microparticles, and the fourth fluorescent image detects the plurality of third microparticles.Anti-Analyte Capture Antibody

[0275] Optionally, prior to contacting the sample with the at least one first capture antibody, the at least one first capture antibody can be bound to a solid support (e.g., a microparticle) which facilitates the separation the first antibody-analyte complex from the sample. Any solid support known in the art can be used, including but not limited to, solid supports made out of polymeric materials in the forms of wells, tubes, or beads (such as a microparticle). The antibody (or antibodies) can be bound to the solid support by adsorption, by covalent bonding using a chemical coupling agent or by other means known in the art, provided that such binding does not interfere with the ability of the antibody to bind analyte. Moreover, if necessary, the solid support can be derivatized to allow reactivity with various functional groups on the antibody. Such derivatization requires the use of certain coupling agents such as, but not limited to, maleic anhydride, N- hydroxysuccinimide and l-ethyl-3-(3-dimethylaminopropyl)carbodiimide.

[0276] After the sample suspected of containing analyte is incubated in order to allow for the formation of a first capture antibody (or multiple antibody)-analyte complex. The incubation can be carried out at a pH of from about 4.5 to about 10.0, at a temperature of from about 2°C to about 45°C, and for a period from at least about one (1) minute to about eighteen (18) minutes, from about 2-6 minutes, from about 7 -12 minutes, from about 5-15 minutes, or from about 3-4 minutes.Detection antibody

[0277] After formation of the first / multiple capture antibody-analyte complex, the complex is then contacted with at least one second detection antibody (under conditions that allow for the formation of a first / multiple antibody-analyte antigen-second antibody complex). In some embodiments, the sample is contacted with the detection antibody simultaneously with the capture antibody. If the first antibody-analyte complex is contacted with more than one detection antibody, then a first / multiple capture antibody-analyte-multiple antibody detection complex is formed. As with first antibody, when the at least second (and subsequent) antibody is brought into contact with the first antibody-analyte complex, a period of incubation under conditions similar to those described above is required for the formation of the first / multiple antibody- analyte-second / multiple antibody complex. Preferably, at least one second antibody contains a detectable label. The detectable label can be bound to the at least one second antibody prior to, simultaneously with or after the formation of the first / multiple antibody-analyte-second / multiple antibody complex. Any detectable label known in the art can be used.

[0278] Chemiluminescent assays can be performed in accordance with the methods described inAdamczyk et al., Anal. Chim. Acta 579(1): 61-67 (2006). Desirably, the formation of pseudobasesin neutral or basic solutions employing an acridinium aryl ester is avoided, such as by acidification. The chemiluminescent response is then recorded. In this regard, the time for recording the chemiluminescent response will depend, in part, on the delay between the addition of the reagents and the particular acridinium employed.

[0279] The order in which the sample and the specific binding partner(s) are added to form the mixture for chemiluminescent assay is not critical. If the first specific binding partner is detectably labeled with an acridinium compound, detectably labeled first specific binding partner-antigen complexes form. Alternatively, if a second specific binding partner is used and the second specific binding partner is detectably labeled with an acridinium compound, detectably labeled first specific binding partner-analyte-second specific binding partner complexes form. Any unbound specific binding partner, whether labeled or unlabeled, can be removed from the mixture using any technique known in the art, such as washing.

[0280] Hydrogen peroxide can be generated in situ in the mixture or provided or supplied to the mixture before, simultaneously with, or after the addition of an above-described acridinium compound. Hydrogen peroxide can be generated in situ in a number of ways such as would be apparent to one skilled in the art.

[0281] Alternatively, a source of hydrogen peroxide can be simply added to the mixture. For example, the source of the hydrogen peroxide can be one or more buffers or other solutions that are known to contain hydrogen peroxide. In this regard, a solution of hydrogen peroxide can simply be added.

[0282] Upon the simultaneous or subsequent addition of at least one basic solution to the sample, a detectable signal, namely, a chemiluminescent signal, indicative of the presence of analyte is generated. The basic solution contains at least one base and has a pH greater than or equal to 10, preferably, greater than or equal to 12. Examples of basic solutions include, but are not limited to, sodium hydroxide, potassium hydroxide, calcium hydroxide, ammonium hydroxide, magnesium hydroxide, sodium carbonate, sodium bicarbonate, calcium hydroxide, calcium carbonate, and calcium bicarbonate. The amount of basic solution added to the sample depends on the concentration of the basic solution. Based on the concentration of the basic solution used, one skilled in the art can easily determine the amount of basic solution to add to the sample. Other labels other than chemiluminescent labels can be employed. For instance, enzymatic labels (including but not limited to alkaline phosphatase) can be employed.

[0283] The chemiluminescent signal, or other signal, that is generated can be detected using routine techniques known to those skilled in the art. Based on the intensity of the signal generated, the amount of analyte of interest in the sample can be quantified. Specifically, the amount of analyte in the sample is proportional to the intensity of the signal generated. The amount of analytepresent can be quantified by comparing the amount of light generated to a standard curve for analyte or by comparison to a reference standard. The standard curve can be generated using serial dilutions or solutions of known concentrations of analyte by mass spectroscopy, gravimetric methods, and other techniques known in the art.Forward Competitive Inhibition

[0284] In a forward competitive format, an aliquot of labeled analyte of interest (e.g., target analyte) having a fluorescent label, a tag attached with a cleavable linker, etc.) of a known amount is used to compete with analyte of interest in a sample for binding to analyte of interest antibody.

[0285] In a forward competition assay, an immobilized specific binding partner (such as an antibody) can either be sequentially or simultaneously contacted with the sample and a labeled analyte of interest, analyte of interest fragment or analyte of interest variant thereof. The analyte of interest peptide, analyte of interest fragment or analyte of interest variant can be labeled with any detectable label, including a detectable label comprised of tag attached with a cleavable linker. In this assay, the antibody can be immobilized on to a solid support. Alternatively, the antibody can be coupled to an antibody, such as an anti species antibody, that has been immobilized on a solid support, such as a microparticle or planar substrate.

[0286] The labeled analyte of interest, the sample and the antibody are incubated under conditions similar to those described above in connection with the sandwich assay format. Two different species of antibody-analyte of interest complexes may then be generated. Specifically, one of the antibody-analyte of interest complexes generated contains a detectable label (e.g., a fluorescent label, etc.) while the other antibody-analyte of interest complex does not contain a detectable label. The antibody-analyte of interest complex can be, but does not have to be, separated from the remainder of the sample prior to quantification of the detectable label. Regardless of whether the antibody-analyte of interest complex is separated from the remainder of the sample, the amount of detectable label in the antibody-analyte of interest complex is then quantified. The amount of analyte of interest (such as membrane-associated analyte of interest, soluble analyte of interest, fragments of soluble analyte of interest, variants of analyte of interest (membrane-associated or soluble analyte of interest) or any combinations thereof) in the sample can then be determined, e.g., as described above.Reverse competition assay

[0287] Tn a reverse competition assay, an immobilized analyte of interest can either be sequentially or simultaneously contacted with a sample and at least one labeled antibody.

[0288] The analyte of interest can be bound to a solid support, such as the solid supports discussed above in connection with the sandwich assay format.

[0289] The immobilized analyte of interest, sample and at least one labeled antibody are incubated under conditions similar to those described above in connection with the sandwich assay format. Two different species analyte of interest-antibody complexes are then generated. Specifically, one of the analyte of interest-antibody complexes generated is immobilized and contains a detectable label (e.g., a fluorescent label, etc.) while the other analyte of interest-antibody complex is not immobilized and contains a detectable label. The non-immobilized analyte of interestantibody complex and the remainder of the sample are removed from the presence of the immobilized analyte of interest-antibody complex through techniques known in the art, such as washing. Once the non-immobilized analyte of interest antibody complex is removed, the amount of detectable label in the immobilized analyte of interest-antibody complex is then quantified following cleavage of the tag. The amount of analyte of interest in the sample can then be determined by comparing the quantity of detectable label as described above.One-Step Immunoassay or "Capture on the Fly"

[0290] In a capture on the fly immunoassay, a solid substrate is pre-coated with an immobilization agent. The capture agent, the analyte and the detection agent are added to the solid substrate together, followed by a wash step prior to detection. The capture agent can bind the analyte and comprises a ligand for an immobilization agent. The capture agent and the detection agents may be antibodies or any other moiety capable of capture or detection as described herein or known in the art. The ligand may comprise a peptide tag and an immobilization agent may comprise an anti-peptide tag antibody. Alternately, the ligand and the immobilization agent may be any pair of agents capable of binding together so as to be employed for a capture on the fly assay (e.g., specific binding pair, and others such as are known in the art). More than one analyte may be measured. In some embodiments, the solid substrate may be coated with an antigen and the analyte to be analyzed is an antibody.

[0291] In certain other embodiments, in a one-step immunoassay or “capture on the fly”, a solid support (such as a microparticle) pre -coated with an immobilization agent (such as biotin, streptavidin, etc.) and at least a first specific binding member and a second specific binding member (which function as capture and detection reagents, respectively) are used. The first specific binding member comprises a ligand for the immobilization agent (for example, if the immobilization agent on the solid support is streptavidin, the ligand on the first specific binding member may be biotin) and also binds to the analyte of interest. The second specific binding member comprises a detectable label and binds to an analyte of interest. The solid support andthe first and second specific binding members may be added to a sample (either sequentially or simultaneously). The ligand on the first specific binding member binds to the immobilization agent on the solid support to form a solid support / first specific binding member complex. Any analyte of interest present in the sample binds to the solid support / first specific binding member complex to form a solid support / first specific binding member / analyte complex. The second specific binding member binds to the solid support / first specific binding member / analyte complex and the detectable label is detected. An optional wash step may be employed before the detection. In certain embodiments, in a one-step assay more than one analyte may be measured. In certain other embodiments, more than two specific binding members can be employed. In certain other embodiments, multiple detectable labels can be added. In certain other embodiments, multiple analytes of interest can be detected, or their amounts, levels or amounts, measured, determined or assessed.

[0292] The use of a capture on the fly assay can be done in a variety of formats as described herein, and known in the art. For example, the format can be a sandwich assay such as described above, but alternately can be a competition assay, can employ a single specific binding member, or use other variations such as are known.Controls

[0293] It may be desirable to include a control sample. The control sample may be analyzed concurrently with the sample from the subject as described above. The results obtained from the subject sample can be compared to the results obtained from the control sample. Standard curves may be provided, with which assay results for the sample may be compared. Such standard curves present levels of marker as a function of assay units, i.e., fluorescent signal intensity, if a fluorescent label is used. Using samples taken from multiple donors, standard curves can be provided for reference levels of the analyte in normal healthy tissue, as well as for “at-risk” levels of the analyte in tissue taken from donors, who may have one or more of the characteristics set forth above.

[0294] Thus, in view of the above, a method for determining the presence, amount, or amount of analyte in a sample is provided. The method comprises assaying the sample for analyte by an immunoassay, for example, employing at least one capture antibody that binds to an epitope on analyte and at least one detection antibody that binds to an epitope on analyte which is different from the epitope for the capture antibody and optionally includes a detectable label, and comprising comparing a signal generated by the detectable label as a direct or indirect indication of the presence, amount or concentration of analyte in the sample to a signal generated as a direct or indirect indication of the presence, amount or concentration of analyte in a calibrator. Thecalibrator is optionally, and is preferably, part of a series of calibrators in which each of the calibrators differs from the other calibrators in the series by the concentration of the analyte.Nucleic Acid analyses

[0295] Various amplification methods and components will be known to one of ordinary skill in the art and any convenient method can be used in the automated assay processing units and methods disclosed herein (see, e.g., Zanoli and Spoto, Biosensors (Basel). 2013 Mar; 3(1): 18-43; Gill and Ghaemi, Nucleosides, Nucleotides, and Nucleic Acids, 2008, 27: 224-243; Craw and Balachandrana, Lab Chip, 2012, 12, 2469-2486; which are herein incorporated by reference in their entirety). Nucleic acid amplification can comprise polymerase chain reaction (PCR), reverse transcription PCR (RT-PCR), quantitative PCR (qPCR), reverse transcription qPCR (RT-qPCR), nested PCR, multiplex PCR, asymmetric PCR, touchdown PCR, random primer PCR, heminested PCR, polymerase cycling assembly (PCA), colony PCR, ligase chain reaction (LCR), digital PCR, methylation specific-PCR (MSP).co-amplification at lower denaturation temperature-PCR (COLD-PCR), allele-specific PCR, intersequence-specific PCR (ISS-PCR), whole genome amplification (WGA), inverse PCR, and thermal asymmetric interlaced PCR (TAIL-PCR).

[0296] Typically, nucleic acid amplification is employed to increase the number of a target nucleic acid (i.e., a target analyte) in the sample, e.g., to thereby facilitate detection of the target nucleic acid. As embodied herein, the nucleic acid amplification methods and system components can be configured to amplify a target nucleic acid using any of a variety or combination of suitable amplification techniques.

[0297] As further disclosed herein, after the sample is prepared, e.g., nucleic acids are isolated from the sample, the isolated nucleic acids can be amplified. The automated assay processing units and methods include contacting the isolated nucleic acids with the amplification oligonucleotides, e.g., forward and reverse primer oligonucleotides, and probes as described herein to fonn a reaction mixture. The reaction mixture is then placed under amplification conditions. The term "amplification conditions," as used herein, refers to conditions that promote annealing and / or extension of the amplification oligonucleotides. In certain embodiments, such conditions include contacting the isolated nucleic acids with an "E-Mix" or a "Core Mix". Typically, an E-Mix is a solution comprising ATP, Phosphocreatine, and buffer. In contrast, a Core Mix typically comprises a collection of proteins necessary to amplify a nucleic acid target.In certain embodiments, such conditions include contacting the isolated nucleic acids with a "MasterMix." As used herein, a MasterMix refers to a solution comprising all of the components, e.g., nucleotide triphosphates, polymerases, primers, and probes, necessary to amplify a target nucleic acid forsubsequent detection, except an activator, which can be separately provided to initiate amplification. For example, an activator is initially dispensed or merged with the sample, the nucleic acid may be isolated with microparticles and the Master Mix is dispensed or merged with the sample. Amplification conditions are well-known in the art and depend on the amplification method selected. In accordance with the disclosed subject matter, amplification conditions encompass a wide range of reaction conditions including, but not limited to, temperature and / or temperature cycling, buffer, salt, ionic strength, pH, and the like.

[0298] Additionally, or alternatively, and in accordance with another aspect of the disclosed subject matter, the automated assay processing units, and methods of the present disclosure include the use of rapid amplification strategies having a duration of about 1 minute to about 60 minutes, about 5 minutes to about 60 minutes, about 8 minutes to about 60 minutes, or in about 8 minutes to about 50 minutes, or in about 8 minutes to about 40 minutes, or in about 8 minutes to about 35 minutes, or in about 8 minutes to about 30 minutes, or in about 8 minutes to about 25 minutes, or about 8 minutes to about 20 minutes, about 1 minute to about 22 minutes, about 5 minutes to about 22 minutes, about 8 minutes to about 22 minutes, about 1 minute to about 20 minutes, about 5 minutes to about 20 minutes, about 8 minutes to about 20 minutes, or about 8 minutes to about 15 minutes from the addition of the reagents sufficient to initiate amplification of a sample of eluted nucleic acids if the targeted nucleic acid(s) is present.

[0299] In certain embodiments, the automated assay processing units, and methods employ isothermal target amplification to amplify target nucleic acid sequences for detection by a automated assay processing unit of the present disclosure. Isothermal target amplification methods do not require a thermocycler, and can be easily adapted and integrated into the automated assay processing units of the present disclosure. As used interchangeably herein, “isothermal amplification reaction”, “isothermal target amplification” and other variations, refers to a target amplification reaction, wherein the temperature does not significantly change during the reaction, i.e., the target amplification reaction is carried out substantially at a single temperature. The temperature of an isothermal amplification reaction does not change over the course of the reaction by more than, e.g., about I0°C, about 9°C, about 8°C, about 7°C, about 6°C, about 5°C, about 4°C, about 3°C, about 2°C, about 1°C.

[0300] Depending on the method of isothermal amplification of nucleic acids, different enzymes are required for the amplification reaction. Known isothermal methods for amplification of nucleic acids are e.g., helicase-dependent amplification (HD A) (Vincent et al, EMBO reports, 2004. 5(8): 795-800), thermostable HDA (tHDA) (An et al, I. Biol. Chem, 2005. 280(32): 28952-28958), strand displacement amplification (SDA) (Walker et al, Nucleic Acids Res, 1992. 20(7): 1691-6), multiple displacement amplification (MDA) (Dean et al. Proc. Natl. Acad. Sci., 2002. 99(8): 5261-5266), rolling circle amplification (Liu et al, J. Am. Chem. Soc., 1996 118: 1587-1594), single primer isothermal amplification (SPIA) (Dafforn et al, Biotechniques, 2004. 37(5):854-7), restriction aided RCA (Wang et al, Genome Res., 2004. 14:2357-2366), transcription mediated amplification (TMA) (Vuorinen et al, J. Clin. Microbiol., 1995. 33:1856-1859), Nucleic Acid Sequence Based Amplification (NASBA) (Kievits et al, J. Virol. Methods, 1991. 35:273-286) and amplification reactions using nicking enzymes, e.g., nicking enzyme amplification reaction (NEAR) (U.S. Patent Application No. 2009 / 017453), amplification reactions using recombination proteins, e.g., recombinase polymerase amplification (RPA) (Piepenburg et al, PLoS Biol., 2004. 4(7): e204), and Loop-mediated isothermal amplification (LAMP) (Notomi et al, Nucleic Acids Res., 2000. 28(12): e63) wherein the at least one mesophilic enzyme for amplifying nucleic acids under isothermal conditions is selected from the group consisting of helicase, mesophilic polymerases, mesophilic polymerases having strand displacement activity, nicking enzymes, recombination proteins, ligases, glycosylases and nucleases.

[0301] In some embodiments, amplification of a target nucleic acid for subsequent detection on the automated assay processing units of the present disclosure is achieved by recombinase polymerase amplification (RPA) (see, U.S. Patent Nos. 7,270,981 ; 7,485,428 and 8,460,875 herein incorporated by reference). RPA is a single tube isothermal amplification reaction. In some cases, reverse transcriptase can be added to an RPA reaction in order to amplify RNA targets. RPA methods employ three enzymes: a recombinase, a single-stranded DNA binding protein (e.g., E. coli SSB) and a strand-displacing polymerase. Generally: first, a recombinase agent is contacted with a first and a second nucleic acid primer to form a first and a second nucleoprotein primer. Second, the first and second nucleoprotein primers are contacted to a double stranded target nucleic acid sequence to form a first double stranded structure at a first portion of said first strand and form a double stranded structure at a second portion of said second strand so the 3’ ends of said first nucleic acid primer and said second nucleic acid primer are oriented towards each other on a given template nucleic acid molecule. Third, the 3’ end of said first and second nucleoprotein primers are extended by a strand-displacing polymerase to generate first and second double stranded nucleic acids, and first and second displaced strands of nucleic acid. The second and third steps are repeated until a desired degree of amplification is achieved. A person skilled in the art will be able to recognize and carry out variations on the general RPA method as described above.

[0302] A recombinase agent is an enzyme that can coat single-stranded DNA (ssDNA) to form filaments, which can then scan double-stranded DNA (dsDNA) for regions of sequence homology. When homologous sequences are located, the nucleoprotein filament (comprising the recombinase agent) strand invades the dsDNA creating a short hybrid and a displaced strand bubble known as a D-loop. Suitable recombinase agents include the E. coli RecA protein or any homologous proteinor protein complex from any phyla. These RecA homologues are generally named Rad51 after the first member of this group to be identified. Other recombinase agents may be utilized in place of RecA, for example, RecT or RecO. Recombinase agents generally require the presence of ATP, ATPyS, or other nucleoside triphosphates and their analogs. Recombinase agents are commonly used in a reaction environment in which regeneration of targeting sites can occur shortly following a round of D-loop stimulated synthesis. This will avoid a stalling of amplification or inefficient linear amplification of ssDNA caused by oscillating single-sided synthesis from one end to the other.

[0303] In some embodiments, amplification of an analyte for subsequent detection the systems and automated assay processing units of the present disclosure is achieved by loop-mediated isothermal amplification (LAMP). LAMP is described in U.S. Patent No. 6,410,278, herein incorporated by reference. Generally, LAMP uses 4-6 primers recognizing 6-8 distinct regions of the target nucleic acid. A strand-displacing DNA polymerase initiates synthesis and two of the primers form loop structures to facilitate subsequent rounds of amplification. A person skilled in the art will be able to recognize and carry out variations on the general LAMP method as described above.

[0304] In some embodiments, amplification of an analyte for subsequent detection on the system and automated assay processing units of the present disclosure is achieved by helicase-dependent amplification (HD A). HDA is based on the unwinding activity of a DNA helicase. HDA relies on one or more helicases to separate (melt, or unwind) two strands of a target nucleic acid duplex. HDA further utilizes a DNA or RNA polymerase to extend primers which are hybridized to single stranded nucleotide sequences to form complementary primer extension products. This process repeats itself so that exponential amplification can be achieved at a single temperature. A person skilled in the art will be able to recognize and carry out variations on the general HDA method as described above. “Complementary” as used herein refers to the complementarity between two nucleic acids, e.g., two DNA molecules. When a nucleotide position in both of the molecules is occupied by nucleotides normally capable of base pairing with each other, then the nucleic acids are considered to be complementary to each other at this position. Thus, two nucleic acids are complementary to each other when a substantial number (at least 50%) of corresponding positions in each of the molecules are occupied by nucleotides which normally base pair with each other (e.g., A:T and G:C nucleotide pairs).

[0305] “Helicase” as used herein refers to any enzyme capable of enzymatically unwinding a double stranded nucleic acid. Any helicase that translocates along DNA or RNA in a 5' to 3' direction or in the opposite 3' to 5' direction may be used. This includes helicases obtained from prokaryotes, viruses, archaea, and eukaryotes or recombinant forms of naturally occurringenzymes as well as analogues or derivatives having the specified activity. Examples of naturally occurring DNA helicases, described by Kornberg and Baker in chapter 1 1 of their book, DNA Replication, W. H. Freeman and Company (2nd ed. (1 92)), include E. coli helicase I, II, III, & IV, Rep, DnaB, PriA, PcrA, T4 Gp41helicase, T4 Dda helicase, T7 Gp4 helicases, SV40 Large T antigen, yeast RAD. Additional helicases that may be useful in HDA include RecQ helicase (Harmon and Kowalczykowski, J. Biol. Chem., 2001. 276:232-243), thermostable UvrD helicases from T. tengcongensis and T. thermophilus (Collins and McCarthy, Extremophiles. 2003, 7:35- 41), thermostable DnaB helicase from T. aquaticus (Kaplan and Steitz, J. Biol. Chem., 1999. 274:6889-6897), and MCM helicase from archaeal and eukaryotic organisms (Grainge et al, Nucleic Acids Res., 2003. 31 :4888-4898).

[0306] In certain embodiments, analyte, e.g., isolated total nucleic acid and / or enriched target nucleic acid, is segregated to a location in which target amplification occurs, e.g., the sample detection zones disclosed above or throughout the present disclosure. This may be done manually, or through automated methods, e.g., through the automated assay processing units and methods disclosed herein.

[0307] In certain embodiments, the steps of target amplification of an analyte are carried out by the automated assay processing units and methods of the present disclosure. In some cases, these steps are carried out by the automated assay processing units of the present disclosures, at temperatures that allow for target amplification. Generally, these automated systems perform steps comprising: i) segregating isolated and / or enriched target nucleic acid, ii) adding enzymes and labelled primers or probes to a sample detection region, iii) adding activators if necessary for target amplification, iv) heating the reaction mixture and v) optionally, quenching the amplification reaction. The automated assay processing units of the present disclosure that carry out the steps of target amplification in an isothermal target amplification reaction are substantially held at a single temperature, depending on the optimal temperature enzymes of various isothermal amplification reactions operate at. In some cases, target amplification carried out by the automated assay processing units of the present disclosure can employ the RPA method, wherein the systems and automated assay processing units are held at substantially a single temperature, e.g., at about 37°C, e.g., about 35-37°C, about 37-39°C, about 36-38°C, about 35-39°C, about 32-42°C. For other isothermal methods, the systems and automated assay processing units are held at a substantially single temperature, e.g., at a temperature of 40°C, or higher, e.g., about 50°C -70°C, such as about 60°C - 65°C. In some cases, target amplification of a target nucleic acid sequence is carried out within 30 min, e.g., in about 25 min, in about 35 min, in about 27 min, in about 29 min, in about 31 min, in about 33 min, in about in at least 10 min e.g., in about 15 min or more, in about 20 min or more, in about 30 min or more.

[0308] In some cases, amplification of a target nucleic acid sequence is performed for a period of 30 min or less, e.g., about 5 min, about 10 min, about 15 min, about 20 min, about or 25 min, about e.g., about 5 min-30 min, about 5 min-25 min, about 5 min-20 min, about 5 min-25 min, about 5 min- 10 min, about 10 min-30 min, about 10 min-25 min, about 10 min-20 min, about 15 min-30 min, about 15 min-25 min, about or about 15 min-20 min.

[0309] In some embodiments, amplification of a target nucleic acid is performed by isothermal amplification, e.g., LAMP, where the amplification is performed for 10 min or less, e.g., about 9 min, about 8 min, about 7 min, about 6 min, about 5 min, about 4 min, about 3min, about 2 min, about 1 min, about 30 sec, about 15 sec, about or less, about e.g., about 15 sec- 10 min, about 30 sec- 10 min, about 1 min- 10 min, about 1 min-5 min, or about 5 min- 10 min.

[0310] In certain embodiments, the automated assay processing units and methods disclosed herein detect a target nucleic acid present at a concentration as low as 1 aM. In certain embodiments, the the systems, automated assay processing units, and methods disclosed herein detect a target nucleic acid present at a concentration of at least 1 aM or more, e.g., about 10 aM, about 100 aM, about 1 fM, about 10 fM, about 100 fM, about 1 pM, about 10 pM, about 100 pM, or more. In certain embodiments, the automated assay processing units and methods disclosed herein detect an amplification product generated from an analyte, where the amplification product is present at a concentration of 25 aM-10 fM. Thus, the presently disclosed detection methods can detect an analyte present in a sample at a concentration of less than 1 aM, prior to amplification. In most instances, an amplification of only 15 times, 10 times, 5 times, 2 times or no amplification is needed to provide sufficient quantities of the target nucleic acid for detection.[0031 1 ] In certain embodiments, the disclosed automated assay processing units and methods can detect an amplification product produced from increasing copy number of the analyte by amplification, where as low as 1000 molecules of the amplification product are produced. Thus, in certain embodiments, a method for detecting presence of a target nucleic acid in a fluid sample may include amplifying the analyte in the sample by amplification to generate as low as 1000 molecules of an amplification product, wherein the amplifying incorporates a tag into the amplification product; capturing the amplification product on a plurality of capture objects comprising a binding member that specifically binds to the tag thereby generating a complex comprising capture object-amplification product; detectably labeling the amplification product in the complex to generate a detectably labeled complex; spatially segregating the capture objects into a plurality of wells such that each well contains no more than one capture object; and detecting the presence of the detectably labeled complex in the plurality of wells.

[0312] The primers or nucleotides used for the amplification reaction may be include a tag, such as, a hapten for incorporation of the tag into the amplified target nucleic acid. Any suitable tagmay be utilized. For example, the tag may be a hapten for which a binding member that specifically binds to the hapten is available. For example, the hapten may be a small molecule for which antibodies that specifically bind to the hapten are available. Exemplary haptens include, avidin, biotin, digoxygenin, dinitrophenyl, dansyl-X, and derivatives thereof. In certain embodiments, the hapten may not be directly optically detectable, i.e., the hapten may not be a dye or a fluorescent molecule because such a hapten may interfere with the digital counting.

[0313] In accordance with the disclosed subject matter, and as embodied herein, the automated assay processing units and methods disclosed herein for analyte can include isothermal amplification methods that rely on nicking and extension amplification reactions (NEAR) to amplify shorter sequences in a quicker timeframe than traditional amplification reactions. These methods can include, for example, reactions that use only two amplification oligonucleotides, one or two nicking enzymes, and a polymerase, under isothermal conditions.

[0314] Typically, in nicking and extension amplification, a target nucleic acid sequence, having a sense and antisense strand, is contacted with a pair of amplification oligonucleotides. The first amplification oligonucleotide comprises a nucleic acid sequence comprising a recognition region at the 3' end that is complementary to the 3' end of the target sequence anti sense strand, a nicking enzyme site upstream of said recognition region, and a stabilizing region upstream of said nicking enzyme site (See, e.g., U.S. Patent Nos 9,689,031; 9,617,586; 9,562,264; and 9,562,263, each of which is incorporated herein by reference in its entirety). The second amplification oligonucleotide comprises a nucleotide sequence comprising a recognition region at the 3' end that is complementary to the 3' end of the target sequence sense strand, a nicking enzyme site upstream of said recognition region, and a stabilizing region upstream of said nicking enzyme site. Two nicking enzymes are provided. One nicking enzyme is capable of nicking at the nicking enzyme site of the first amplification oligonucleotide but incapable of nicking within said target sequence. The other nicking enzyme is capable of nicking at the nicking enzyme site of the second amplification oligonucleotide but incapable of nicking within said target sequence. A DNA polymerase is employed under conditions for amplification which involves multiple cycles of extension of the amplification oligonucleotides thereby producing a double-stranded nicking enzyme site which are nicked by the nicking enzymes to produce the amplification product. For example, see U.S. Patent Nos: 9,689,031; 9,617,586; 9,562,264; 9,562,263; and 10,851,406, U.S. Patent Application Publication No. 2018 / 0023130 and U.S. Patent Application No.: 16 / 243 / 829, each of which is incorporated herein by reference in its entirety.

[0315] Tn some embodiments, reactions use only two templates to prime, one or two nicking enzymes, and a polymerase, under isothermal conditions. In exemplary embodiments, the polymerase and the nicking enzyme are thermophilic, and the reaction temperature is significantlyabove the melting temperature of the hybridized target region. The nicking enzyme nicks only one strand in a double-stranded duplex, so that incorporation of modified nucleotides is not necessary as it is in strand displacement. In some embodiments, the method is able to amplify RNA without a separate reverse transcription step, although conversion of RNA to DNA by reverse transcription may be used if desired.

[0316] In some embodiments, the method comprises contacting a target DNA molecule comprising a double-stranded target sequence having a sense strand and an antisense strand, with a forward template and a reverse template, wherein said forward template comprises a nucleic acid sequence comprising a recognition region at the 3' end that is complementary to the 3' end of the target sequence anti sense strand; a nicking enzyme site upstream of said recognition region, and a stabilizing region upstream of said nicking enzyme site; the reverse template comprises a nucleotide sequence comprising a recognition region at the 3' end that is complementary to the 3' end of the target sequence sense strand, a nicking enzyme site upstream of the recognition region, and a stabilizing region upstream of the nicking enzyme site; providing a first nicking enzyme that is capable of nicking at the nicking enzyme site of the forward template, and does not nick within the target sequence; providing a second nicking enzyme that is capable of nicking at the nicking enzyme site of the reverse template and does not nick within the target sequence; and providing a DNA polymerase; under conditions wherein amplification is performed by multiple cycles of the polymerase extending the forward and reverse templates along the target sequence producing a double-stranded nicking enzyme site, and the nicking enzymes nicking at the nicking enzyme sites, producing an amplification product.

[0317] In certain embodiments, the DNA polymerase is a thermophilic polymerase. In other examples, the polymerase and said nicking enzymes are stable at temperatures up to about 37°C, about 42°C, about 60°C, about 65°C, about 70°C, about 75°C, about 80°C, or about 85°C. In certain embodiments, the polymerase is stable up to 60°C. The polymerase may, for example, be selected from the group consisting of Bst (large fragment), 9° N, VentR® (exo-) DNA Polymerase, THERMINATOR, and THERMINATOR II (New England Biolabs).

[0318] The nicking enzyme may, for example, nick upstream of the nicking enzyme binding site, or the nicking enzyme may nick downstream of the nicking enzyme binding site. In certain embodiments, the forward and reverse templates comprise nicking enzyme sites recognized by the same nicking enzyme and the first and the second nicking enzyme are the same. The nicking enzyme may, for example, be selected from the group consisting of Nt.BspQI, Nb.BbvCi, Nb.BsmI, Nb.BsrDI, Nb.BtsI, Nt.AlwI, Nt.BbvCI, Nt.BstNBI, Nt.CviPIT, Nb.BpulOI, and Nt.BpulOI.

[0319] In some embodiments, the target sequence comprises from 1 to 5 nucleotides more than the sum of the nucleotides of said forward template recognition region and said reverse template recognition region.

[0320] In some embodiments, the forward template is provided at the same concentration as the reverse template. In other examples, the forward template is provided at a ratio to the reverse template at the range of ratios of 1 : 100 to 100: 1.

[0321] As embodied herein, the NEAR reaction time can be about 10 minutes to about 30 minutes, or about 8 minutes to about 25 minutes, or about 8 minutes to about 20 minutes, or even about 8 minutes to about 15 minutes from the addition of the reagents sufficient to initiate NEAR amplification. In certain embodiments, the NEAR reaction time is about 1 minute to about 20 minutes, about 5 minutes to about 20 minutes, about 8 minutes to about 20 minutes, about 1 minute to about 10 minutes or about 5 minutes to about 10 minutes from the addition of the reagents sufficient to initiate NEAR amplification.

[0322] Provided herein are methods for measuring or detecting an analyte present in a biological sample. In some embodiments, the target analyte is a target nucleic acid. In some embodiments, the target nucleic acid is DNA. In some embodiments, the DNA is single stranded DNA. In some embodiments, the DNA is double stranded DNA. In some embodiments, the target nucleic acid is RNA. In such methods, a consumable comprising wells is loaded into the automated assay processing unit. Following the loading of the consumable, a number of fluids may be dispensed into various regions prior to further processing the sample in an assay. In some embodiments, a sample comprising a target analyte is dispensed into a first region of the consumable. In some embodiments, a particle solution is dispensed into the first region of the consumable. The particle solution may comprise a single type of particle or multiple types of particles. In some embodiments, the particle solution comprises microparticles. In some embodiments, the particle solution comprises microparticles and assisting microparticles. The microparticles are attached to a first specific binding partner that specifically binds to the target nucleic acid. In some embodiments, the specific binding member is a nucleic acid that specifically hybridizes to the target nucleic acid. In some embodiments, reagents in that amplify and detect the target analyte under isothermal amplification conditions comprising a detection probe that produces a detectable signal following amplification of the target analyte are dispensed into a third region. Additionally, wash buffer may be dispensed into at least a third region of the consumable. The wash buffer may be dispensed into more than the third region, such as a fourth, a fifth, a sixth, a seventh, an eighth, a nineth, a tenth, an eleventh, a twelfth, a thirteenth, a fourteenth, a fifteenth, or more than a fifteenth region. Optionally, a lysis buffer may be dispensed into the first region.

[0323] A number of fluids are dispensed into various regions of the consumable. The fluids may be dispensed in any order deemed useful. For instance, the sample, the particle solution, the reagent that amplify and detect the target analyte under isothermal amplification conditions, e.g., the isothermal amplification reagents, the wash buffer, and the lysis buffer may be dispensed at the same time or at different times. In some embodiments, the sample is dispensed from a sample dispenser comprising a sample pump and a sample nozzle. In some embodiments, the isothermal amplification reagents, the wash buffer, and the lysis buffer may be dispensed from one or more reagent dispensers. The isothermal amplification reagents, the wash buffer, and the lysis buffer may be dispensed from the same reagent dispenser or different reagent dispensers.

[0324] Following sample and particle solution dispensing, the sample may be incubated with the microparticles or microparticles and assisting microparticles for a time sufficient to allow binding of the first specific binding partner to an analyte present in the sample thereby producing microparticles bound to the target analyte where the microparticles are attached to the first specific binding partner and the first specific binding partner is hybridized to the target analyte. During the incubation the consumable may be heated to a desired temperature using a heating element. In some embodiments, the desired temperature is 30-45 C. The sample may be mixed with the microparticles or microparticles and assisting microparticles using a mixing component. In some embodiments, mixing comprises engaging the engagement arm of the mixing component with at least a portion of the consumable and vibrating or agitating the consumable. In some embodiments, the mixing comprises contacting the first region with a ballistic mixing device of a mixing component and agitating the first region.

[0325] Optionally, the microparticles bound to the target analyte or microparticles bound to the target analyte and assisting particles may be subjected to a magnetic field to move the microparticles bound to the target analyte to a third region of the consumable comprising wash buffer. In some embodiments, the subjecting the microparticles bound to the target analyte to a magnetic field comprises positioning the first magnet under the first region, un-retracting the first magnet, moving the first magnet or the consumable positioning the first magnet under the thrid region, and retracting the first magnet. Once in the third region, the microparticles or the microparticles and assisting microparticles may be mixed with the wash buffer. In some embodiments, the mixing comprises positioning the second magnet over the fourth region, unretracting the second magnet, and retracting the second magnet.

[0326] Next, the microparticles bound to the detectably labeled target analyte or microparticles bound target analyte and assisting particles may be subjected to a magnetic field to move the microparticles to a second region of the consumable comprising wells. The microparticles bound to the target analyte may be allowed to settle into the array of wells in the second region. Themicroparticles may settle using gravitational force or by applying electric or magnetic force. The wells contain the isothermal amplification reagents. The reagents necessary for the amplification and detection of the target analyte may be any of the reagents discussed above. Suitable reagents include, without limitation, a polymerase, a reverse transcriptase, a strand-displacing polymerase, forward and reverse primers specific to the target analyte, a recombinase, a single-stranded DNA binding protein, dNTPs, at least four primers that recognize at least 6 distinct regions in the target analyte, one or more helicases, a detection probe that produces a detectable signal following amplification of the target analyte, a first amplification oligonucleotide comprising a first nicking enzyme site, a second amplification oligonucleotide comprising a second nicking enzyme site, a first nicking enzyme that recognizes the first nicking enzyme site and second nicking enzyme recognizing the second enzyme site, etc. The reagents may also include reagents that elute the target analyte from the microparticles such that the target analyte is no longer bound to the first specific binding member. Next, the wells containing the microparticles bound to the target analyte are sealed using a hydrophobic liquid (e.g., oil). In some embodiments, the sealing comprises dispensing from a hydrophobic liquid dispenser the hydrophobic liquid into the third region thereby flowing the hydrophobic liquid over the wells. Lastly, the wells are imaged thereby detecting the detectable signal produced by the detection probe.

[0327] In some embodiments, the imaging of the wells comprises i) immobilizing the consumable, ii) adjusting a focus of an imaging component to a first focal plane, iii) capturing a first scatter image, iv) capturing a first fluorescent image, and v) repeating steps ii-iv) one or more times to generate a stack of scatter images and fluorescent images spanning a plurality of focal planes. In some embodiments, step iii) comprises enabling one or more scatter LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more scatter LEDs. In some embodiments, step iv) comprises enabling one or more fluorescent LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more fluorescent LEDs. In some embodiments, step i) comprises contacting an alignment datum with the consumable thereby activating a raising element and a clamping component, wherein: the raising element is positioned below the consumable and the clamping component is positioned above the consumable, and the raising element provides an upward force on the consumable and the clamping component provides a downward force on the consumable.

[0328] In some instances, it may be desirable to analyze multiple analytes at once. In these embodiments, the microparticles comprise a plurality of first microparticles and a plurality of second microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first nucleic acid sequence and the first specific binding partner of the plurality of first microparticles is specific to a second nucleic acid sequence, and the plurality of firstmicroparticles comprise a first fluorophore and the plurality of second microparticles comprise a second fluorophore. In some embodiments, the microparticles comprise a plurality of first microparticles, a plurality of second microparticles, and a plurality of third microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first nucleic acid sequence, the first specific binding partner of the plurality of second microparticles is specific to a second nucleic acid sequence, the first specific binding partner of the plurality of third microparticles is specific to a third nucleic acid sequence, and the plurality of first microparticles comprise a first fluorophore, the plurality of second microparticles comprise a second fluorophore, and the plurality of third microparticles comprise a third fluorophore. . In some embodiments, the first fluorophore, the second fluorophore, and the third fluorophore are the same fluorophore. In some embodiments, the first fluorophore, the second fluorophore, and the third fluorophore are different fluorophores. In some embodiments, the second fluorophore and the third fluorophore emits at the same wavelength but are excited by a different wavelength of light. When two or more of the fluorophores are the same fluorophore, the fluorophores may be present in different concentrations such that the concentration of the first fluorophore is a lower or higher concentration than that of the second fluorophore. In some embodiments, the first nucleic acid sequence, the second nucleic acid sequence, and the third nucleic acid sequence are the same nucleic acid sequence. In some embodiments, the first nucleic acid sequence, the second nucleic acid sequence, and the third nucleic acid sequence are different nucleic acid sequences.

[0329] In some embodiments, the first nucleic acid sequence and the second nucleic acid sequence are the same nucleic acid sequence. In some embodiments, the first nucleic acid sequence and the second nucleic acid sequence are different nucleic acid sequences. When multiple analyte detection is desired, the target analyte is a first target analyte and a second target analyte and the first target analyte is different from the second target analyte. In some embodiments, the target analyte is a first target analyte, a second target analyte, and a third target analyte and the first, second, and third target analyte are different. In some embodiments, the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles. In some embodiments, the plurality of first microparticles are coated with the first fluorophore and the plurality of second microparticles are coated with the second fluorophore. In some embodiments, the plurality of first microparticles are embedded with the first fluorophore and the plurality of second microparticles are embedded with the second fluorophore. In some embodiments, capturing the first fluorescent image of step iv) further comprises capturing a second fluorescent and capturing a third fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, and the third fluorescent image detects the plurality ofsecond microparticles. In some embodiments, the second target analyte is a control analyte. By “control analyte” it is meant that the analyte is used as a control to determine if the assay steps were performed without error and the assay reagents performed as expected. The control analyte may be an analyte that is added to the sample in a known concentration. In an alternative embodiment, the control analyte may be endogenously present in the sample. In some embodiments, the control analyte is biotin. When the control analyte is biotin, the first specific binding partner of the plurality of second microparticles is antibody or fragment thereof that binds to biotin. In some embodiments, the control analyte is biotin. When the control analyte is biotin, the first specific binding partner of the plurality of second microparticles is streptavidin.

[0330] In some embodiments, the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles and / or the plurality of third microparticles. In some embodiments, the plurality of first microparticles are coated with the first fluorophore, the plurality of second microparticles are coated with the second fluorophore, and the plurality of third microparticles are coated with the third fluorophore. In some embodiments, the plurality of first microparticles are embedded with the first fluorophore, the plurality of second microparticles are embedded with the second fluorophore, and the plurality of third microparticles are embedded with the third fluorophore. In some embodiments, capturing the first fluorescent image of step iv) further comprises capturing a second fluorescent and capturing a third fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, and the third fluorescent image detects the plurality of second microparticles. In some embodiments, capturing the first fluorescent image of step iv) further comprises capturing a second fluorescent, capturing a third fluorescent image, and capturing fourth fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, the third fluorescent image detects the plurality of second microparticles, and the fourth fluorescent image detects the plurality of third microparticles.

[0331] In some embodiments, the reagents necessary for the amplification and detection of the target analyte are for RPA. When the reagents are for RPA, the reagents comprise dNTPs, a recombinase, a single-stranded DNA binding protein, a strand-displacing polymerase, a forward and reverse primer specific to the target analyte, and a detection probe that produces a detectable signal following amplification of the target analyte. In some embodiments, the reagents necessary for the amplification and detection of the target analyte are for LAMP. When the reagents are for LAMP, the reagents comprise dNTPs, at least four primers that recognize at least 6 distinct regions in the target analyte, a strand-displacing polymerase, and a detection probe that produces adetectable signal following amplification of the target analyte. In some embodiments, the reagents necessary for the amplification and detection of the target analyte are for HDA. When the reagents are for HDA, the reagents comprise dNTPs, one or more helicases, a forward and reverse primer specific to the target analyte, a polymerase, and a detection probe that produces a detectable signal following amplification of the target analyte. In some embodiments, the reagents necessary for the amplification and detection of the target analyte are for NEAR. When the reagents are for NEAR, the reagents comprise dNTPs, a DNA polymerase, a first amplification oligonucleotide comprising a first nicking enzyme site, a second amplification oligonucleotide comprising a second nicking enzyme site, a first nicking enzyme that recognizes the first nicking enzyme site and second nicking enzyme recognizing the second enzyme site, and a detection probe that produces a detectable signal following amplification of the target analyte.EXAMPLESExample 1

[0332] The throughput of automated assay processing units can be limited by the fact that devices are often designed for one assay test per consumable. One method to increase the throughput of automated assay processing units is to increase the number of tests that can be performed simultaneously on a single consumable. The use of two different colored particles in a single consumable enables two separate assays to be performed simultaneously. These beads differ in color by means of the fluorophore. The fluorophore is either coated or embedded in the material of the particle. Each colored particle is then coated with a protein (i.e., antibody / or antigen) or nucleic acid, which is targeted to a specific analyte. A sample (serum / plasma), which contains a mixture of target analytes is then added to the consumable. The two bead types are then added to the sample and then mixed. The beads capture their targeted analyte in the sample and then are processed through the consumable on the automated assay processing unit. An exemplary automated assay processing unit may have three different LEDs (Green, Blue and Red). The two- colored bead populations can separated using the green and red channels on the automated assay processing unit in a consumable containing a well array. The blue LED can be utilized to determine presence of analyte bound to both types of beads and quantitate analyte present in the sample.

[0333] FIG. 4 discloses an experiment in which a first population of green particles and a second population of red particles were used to detect beta-human chorionic gonadotropin (B-hCG). Green particles coated with anti-B-hCG antibodies were mixed with red particles coated with B- hCG, in a 50 / 50 ratio. The B-hCG calibrator set was tested as samples. Signal separation was present between most of the calibrator levels.

[0334] FIG. 5-17 disclose the results of a multiplex assay where AFP and T-PSA are detected in a sample using green particles that bind T-PSA and red particles that bind AFP. In the assay, the consumable was loaded with 40 mL of substrate solution. The consumable was then transferred to the automated assay processing unit and the wash channels were filled with 25 mL of DEA wash buffer. 15ul of conjugate was mixed for each assay in a centrifuge tube and vortexed. 15 mL of the conjugate was transferred to the conjugate port to get roughly equal amounts for each assay. In a centrifuge tube, 15 mL AFP sample, 20 mL TPS A sample, 605k T-PSA green particles and 358k AFP red particles were added. The centrifuge was vortexed for 18 seconds and placed in a thermomixer for 60 seconds at 1500 RPM and 38C for IR incubation. The contents were transferred from the centrifuge tube into 2-step sample well on the consumable. 255k helper beads were added in the 2-step sample well and proceed to wash. Conjugate incubation is for 60 seconds. The oil seal was completed and the waste was emptied. Took images. Script is setup to take 21 images with green, blue, and red LEDs per stack and 8 stacks total.

[0335] FIG. 5 discloses that non-multiplexed AFP can be detected with low analyte samples using digital mode (up to Calibrator C). The detection range can be extended using analog mode for higher analyte samples (up to Calibrator E). The sample and particles were incubated together for 1 minute. The conjugated with the particles was incubated for 1 minute. The substrate solution was incubated with the particles for 4.5 minutes.

[0336] FIG. 6 discloses the results for AFP when multiplexed with T-PSA (right chart) vs. AFP non-multiplexed (left chart) on the automated assay processing unit. Good signal separation was observed between calibrator levels when AFP was multiplexed with T-PSA. However; the multiplexed active particle signals are reduced across the calibrator set, when compared to the non-multiplexed method.

[0337] FIG. 7 discloses the microparticle seeding for AFP when multiplexed with T-PSA (right chart) vs. AFP non-multiplexed (left chart) on the automated assay processing unit. Red particles were used for AFP non-multiplexed and multiplexed testing.

[0338] FIG. 8 discloses the results for AFP when multiplexed with T-PSA (right chart) vs. AFP non-multiplexed (left chart) on the automated assay processing unit. Good signal separation was observed between low analyte levels when AFP was multiplexed with T-PSA. However; the multiplexed active particle signals are reduced across the low analytes, when compared to the nonmultiplexed method.

[0339] FIG. 9 discloses the microparticle seeding for AFP when multiplexed with T-PSA (right chart) vs. AFP non-multiplexed (left chart) on the automated assay processing unit. Red particles were used for AFP non-multiplexe...

Claims

WHA IS CLAIMED IS:

1. An automated assay processing unit, comprising: a first module comprising a sample processing component comprising a consumable holder; a second module comprising an aspiration and dispense component; a third module comprising a mixing component; and a fourth module comprising an optics component, wherein the sample processing component is configured to translate the consumable holder to a plurality of positions within the sample processing component, the first, second, third, and fourth modules are operably connected and configured to interface with a consumable located on the consumable holder.

2. The automated assay processing unit of claim 1, wherein the sample processing component comprises a first retractable magnet beneath the consumable holder that is configured to be proximate to the consumable holder in an un-retracted state and distal to the consumable holder in a retracted state.

3. The automated assay processing unit of claim 2, wherein the first retractable magnet is encased in a sleeve in the retracted state wherein the sleeve shields the first retractable magnet thereby reducing the local magnetic field.

4. The automated assay processing unit of any of claims 1-3, wherein the plurality of positions comprise a first position, a second position, and a third position.

5. The automated assay processing unit of claim 4, wherein the sample processing component comprises one or more compression elements that are configured to engage the consumable holder, wherein the consumable holder is in the second position.

6. The automated assay processing unit of claims 4 or 5, wherein the sample processing component comprises a first heating element located in the first position beneath the consumable holder and configured to heat a consumable present in the consumable holder to a first temperature, wherein the consumable holder is positioned in the first position.

7. The automated assay processing unit of any of claims 4-6, wherein the sample processing component comprises a second heating element located in the second position beneath the consumable holder and configured to heat a consumable present in the consumable holder to a second temperature, wherein the consumable holder is positioned in the second position.

8. The automated assay processing unit of claims 6 or 7, wherein the first temperature is different from the second temperature.

9. The automated assay processing unit of claims 6 or 7, wherein the first temperature is same as the second temperature.

10. The automated assay processing unit of any of claims 4-9, wherein the sample processing component comprises a first temperature sensor in the first position.1 1. The automated assay processing unit of any of claims 4-10, wherein the sample processing component comprises a second temperature sensor in the second position.

12. The automated assay processing unit of any of claims 4-11, wherein the sample processing component comprises a raising element in the third position configured to raise the consumable holder and engage the consumable holder with the optics component, wherein the consumable holder is in the third position.

13. The automated assay processing unit of claim 12, wherein the raising element raises the consumable holder when the consumable holder physically contacts the optics component.

14. The automated assay processing unit of claims 12 or 13, wherein the raising element is an imaging component of the optics component.

15. The automated assay processing unit of claim 13, wherein the consumable physically contacts an alignment datum present on the optics component.

16. The automated assay processing of any of claims 1-15, wherein the aspiration and dispense component is positioned above the sample processing component.

17. The automated assay processing unit of any of claims 1-16, wherein the aspiration and dispense component comprises an aspiration nozzle fluidically connected to an aspiration pump.

18. The automated assay processing unit of claims 17, wherein the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle when the consumable holder is in the second position.

19. The automated assay processing unit of any of claims 1-18, wherein the aspiration and dispense component comprises a hydrophilic liquid nozzle fluidically connected to a hydrophilic liquid pump.

20. The automated assay processing unit of claim 19, wherein the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle, wherein the consumable holder is in the second position.

21. The automated assay processing unit of any of claims 1-20, wherein the aspiration and dispense component comprises a hydrophobic liquid nozzle fluidically connected to a hydrophobic liquid pump.

22. The automated assay processing unit of claim 21, wherein the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle when the consumable holder is in the second position.

23. The automated assay processing unit of claim 22, wherein the aspiration pump is configured to aspirate the hydrophilic liquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously when the consumable holder is in the second position.

24. The automated assay processing unit of any of claims 1-23, wherein the aspiration and dispense component comprises a second retractable magnet.

25. The automated assay processing unit of claim 24, wherein the second retractable magnet is positioned above the consumable holder, wherein the consumable holder is in the secondposition that is configured to be proximate to the consumable holder in an un-retracted state and distal to the consumable holder in a retracted state.

26. The automated assay processing unit of claim 25, wherein the second retracted magnet is positioned in a position that is horizontally offset from the first retractable magnet.

27. The automated assay processing unit of claim 25 or 26, wherein the first retractable magnet and the second retractable magnet are configured to not be in the un-retracted state at the same time.

28. The automated assay processing unit of any of claims 1 -27, wherein the aspiration and dispense component comprises one or more reagent pumps fluidically connected to one or more reagent nozzles.

29. The automated assay processing unit of claim 28, wherein the one or more reagent pumps are configured to dispense a reagent through the one or more reagent nozzles , wherein the consumable holder is in the second position.

30. The automated assay processing unit of any of claims 1-27, wherein the mixing component comprises one or more reagent pumps fluidically connected to one or more reagent nozzles.

31. The automated assay processing unit of claim 30, wherein the one or more reagent pumps are configured to dispense a reagent through the one or more reagent nozzles when the consumable holder is in the second position.

32. The automated assay processing unit of any of claims 1-31, wherein the mixing component comprises a mixing motor and an engagement arm that is configured to engage with a consumable present in the consumable holder.

33. The automated assay processing unit of claim 32, wherein the engagement ami engages with a portion of the consumable.

34. The automated assay processing unit of claim 32, wherein the engagement arm engages with the entire consumable.

35. The automated assay processing unit of any of claims 1-31, wherein the mixing component comprises a ballistic mixing device.

36. The automated assay processing unit of claim 35, wherein the ballistic mixing device is a pipettor.

37. The automated assay processing unit of any of claims 1-36, wherein the optics component comprises imaging component and a secondary illumination component.

38. The automated assay processing unit of claim 37, wherein the imaging component is positioned beneath the consumable holder, wherein the consumable holder is in the third position.

39. The automated assay processing unit of claims 37 or 38, wherein the imaging component comprises an illumination component comprising one or more LEDs encased in an enclosed illumination chamber having an opening positioned beneath consumable holder, wherein the consumable holder is in the third position.

40. The automated assay processing unit of claim 39, wherein the one or more LEDs are in proximity to one or more thermistors.

41. The automated assay processing unit of claims 39 or 40, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

42. The automated assay processing unit of claim 41, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

43. The automated assay processing unit of claims 41 or 42, wherein the one or more thermistors are two or more, or three or more thermistors.

44. The automated assay processing unit of claim 43, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

45. The automated assay processing unit of any of claims 39-44, wherein the one or more LEDs are present on a printer circuit board (PCB), wherein the PCB maintains the LEDs, the enclosed illumination chamber, and the consumable holder at a third temperature, wherein the consumable holder is at the third position.

46. The automated assay processing unit of claim 45, wherein the PCB comprises an aluminum core.

47. The automated assay processing unit of any of claims 1-46, wherein the optics component comprises a clamping component.

48. The automated assay processing unit of claim 47, wherein the clamping component engages with the consumable holder and provides a downward clamping force, wherein the consumable holder physically contacts the optics component in the third position.

49. The automated assay processing unit of any of claims 37 -48, wherein the secondary illumination component comprises one or more LEDs positioned beneath the consumable holder thereby enabling capture of a forward scatter image, wherein the consumable holder is in the third position50. The automated assay processing unit of any of claims 37 -48, wherein the secondary illumination component comprises one or more LEDs positioned above the consumable holder thereby enabling capture of a back scatter image, wherein the consumable holder is in the third position51. The automated assay processing unit of claims 49 or 50, wherein the one or more LEDs are in proximity to one or more thermistors.

52. The automated assay processing unit of any of claims 49-51, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

53. The automated assay processing unit of claim 52, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

54. The automated assay processing unit of claims 52 or 53, wherein the one or more thermistors are two or more, or three or more thermistors.

55. The automated assay processing unit of claim 54, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

56. The automated assay processing unit of any of claims 49-54, wherein the one or more LEDs are present on a second printer circuit board (PCB).

57. The automated assay processing unit of claim 56, wherein the second PCB comprises an aluminum core.

58. The automated assay processing unit of any of claims 1-57, wherein the automated assay processing unit capable of being connected to one or more different automated assay processing units.

59. The automated assay processing unit of claim 58, wherein the one or more different automated assay processing units are adjacent to the automated assay processing unit such that the one or more automated assay processing units are parallel to the automated assay processing unit.

60. The automated assay processing unit of claim 58, wherein the one or more different automated assay processing units are in series to the automated assay processing unit such that the one or more automated assay processing units are connected to the automated assay processing unit at the end closest to the first position or the third position.

61. The automated assay processing unit of claim 58, wherein the one or more different automated assay processing units are vertically offset from the automated assay processing unit such that the one or more automated assay processing units are stacked above or below the automated assay processing unit.

62. The automated assay processing unit of any of claims 58-61, wherein the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.

63. The automated assay processing unit of any of claims 1-62, further comprising a consumable.

64. The automated assay processing unit of claim 63, wherein the consumable is a microfluidic device.

65. An automated medium-capacity assay processing unit, comprising: two or more fluid addition stations, two or more wash stations, two or more imaging stations, and a consumable mover.

66. The automated medium-capacity assay processing unit of claim 65, wherein the consumable mover comprises a conveyer system operably coupled to the fluid addition stations, wash stations, and imaging stations wherein the consumable mover is configured to move a consumable from a first fluid addition station of the fluid addition stations to a first wash station of the wash stations and from the first wash station to a first imaging station of the imaging stations.

67. The automated medium-capacity assay processing unit of claim 66, wherein the conveyer system comprises a consumable holder that is capable of being moved between each station.

68. The automated medium-capacity assay processing unit of claim 65, wherein the consumable mover comprises a griping component that is capable of gripping a consumable and moving the consumable between each station.

69. The automated medium-capacity assay processing unit of any of claims 65-68, wherein the two or more fluid addition stations comprise one or more heating elements configured to heat a consumable to a first temperature, wherein the consumable is present in a first fluid addition station of the two or more fluid addition stations.

70. The automated medium-capacity assay processing unit of any of claims 65-69, wherein the two or more fluid addition stations comprise one or more reagent dispensers comprising one or more reagent pumps fluidically connected to one or more reagent nozzles.

71. The automated medium-capacity assay processing unit of claim 70, wherein the two or more fluid addition stations share the one or more reagent dispensers.

72. The automated medium-capacity assay processing unit of claim 70, wherein each fluid addition station comprises a unique reagent dispenser of the one or more reagent dispensers.

73. The automated medium-capacity assay processing unit of any of claims 65-72, wherein the two or more fluid addition stations comprise a hydrophilic liquid dispenser comprising a hydrophilic liquid nozzle fluidically connected to a hydrophilic liquid pump.

74. The automated medium-capacity assay processing unit of claim 73, wherein the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle when the consumable holder is in the fluid addition station.

75. The automated medium-capacity assay processing unit of any of claims 65-74, wherein the two or more fluid addition stations comprise a mixing component.

76. The automated medium-capacity assay processing unit of claim 71, wherein the two or more fluid addition stations share the mixing component.

77. The automated medium-capacity assay processing unit of claim 71, wherein the mixing component is a first mixing component of a plurality of mixing components, and each fluid addition station comprises a dedicated mixing component of the plurality of mixing components.

78. The automated medium-capacity assay processing unit of any of claims 71-77, wherein the mixing component comprises a mixing motor and an engagement arm that is configured to engage with a consumable when the consumable is present in a first fluid addition station of the two or more fluid addition stations79. The automated medium-capacity assay processing unit of claim 78, wherein the engagement arm engages with a portion of the consumable.

80. The automated medium-capacity assay processing unit of claim 78, wherein the engagement arm engages with the entire consumable.

81. The automated medium-capacity assay processing unit of claim 78, wherein the mixing component comprises a ballistic mixing device.

82. The automated medium-capacity assay processing unit of claim 81, wherein the ballistic mixing device is a pipettor.

83. The automated medium-capacity assay processing unit of any of claims 65-82, wherein the two or more wash stations comprise a first retractable magnet positioned beneath a consumable, wherein the consumable is present in the two or more wash stations, wherein the first retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state.

84. The automated medium-capacity assay processing unit of claim 83, wherein the first retractable magnet is encased in a sleeve in the retracted state wherein the sleeve shields the first retractable magnet thereby reducing the local magnetic field.

85. The automated medium-capacity assay processing unit of claim 83 or 84, wherein the two or more wash stations share the first retractable magnet.

86. The automated medium-capacity assay processing unit of claim 83 or 84, wherein each wash station comprises the first retractable magnet.

87. The automated medium-capacity assay processing unit of claims 65-86, wherein the two or more wash stations comprise one or more heating elements configured to heat one or more consumables to a second temperature, wherein the one or more consumables are present in the two or more wash stations.

88. The automated medium-capacity assay processing unit of any of claims 65-87, wherein the two or more wash stations comprise a second retractable magnet.

89. The automated medium-capacity assay processing unit of claim 88, wherein the second retractable magnet is positioned above a consumable, wherein the consumable is in thetwo or more wash stations, wherein the second retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state.

90. The automated medium-capacity assay processing unit of claim 88 or 89, wherein the second retracted magnet is positioned in a position that is horizontally offset from the first retractable magnet.

91. The automated medium-capacity assay processing unit of claim 89 or 90, wherein the first retractable magnet and the second retractable magnet are configured to not be in the unretracted state at the same time.

92. The automated medium-capacity assay processing unit of any of claims 65-91, wherein two or more wash stations comprise an aspiration device comprising an aspiration nozzle fluidically connected to an aspiration pump.

93. The automated medium-capacity assay processing unit of claims 92, wherein the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle, wherein a consumable is present in the two or more wash stations.

94. The automated medium-capacity assay processing unit of claim 92 or 93, wherein the two or more wash stations share the aspiration device.

95. The automated medium-capacity assay processing unit of claim 92 or 93, wherein the aspiration device is a first aspiration device of a plurality of aspiration devices, and each wash station comprises a dedicated aspiration device of the plurality of aspiration devices.

96. The automated medium-capacity assay processing unit of any of claims 65-93, wherein the two or more wash stations comprise a hydrophobic liquid dispenser comprising a hydrophobic liquid nozzle fluidically connected to a hydrophobic liquid pump.

97. The automated medium-capacity assay processing unit of claim 96, wherein the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle, wherein a consumable is present in the two or more wash stations.

98. The automated medium-capacity assay processing unit of claim 97, wherein the aspiration pump is configured to aspirate the hydrophilic liquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously, wherein a consumable is present in the two or more wash stations.

99. The automated medium-capacity assay processing unit of claim 92 or 93, wherein the two or more wash stations share the hydrophobic liquid dispenser.

100. The automated medium-capacity assay processing unit of claim 92 or 93, wherein the hydrophobic liquid dispenser is a first hydrophobic liquid dispenser of a plurality of hydrophobic liquid dispensers, and each wash station comprises a dedicated hydrophobic liquid dispenser of the plurality of hydrophobic liquid dispensers.

101. The automated medium-capacity assay processing unit of any of claims 65-100, wherein the two or more imaging stations comprise an imaging component and a secondary illumination component.

102. The automated medium-capacity assay processing unit of claim 101, wherein the imaging component is positioned beneath a consumable, wherein the consumable present in the two or more imaging stations.

103. The automated medium-capacity assay processing unit of claims 101 or 102, wherein the imaging component comprises an illumination component comprising one or more LEDs encased in an enclosed illumination chamber having an opening positioned beneath the consumable holder, wherein the consumable holder is present in the two or more imaging stations.

104. The automated medium-capacity assay processing unit of claim 103, wherein the one or more LEDs are in proximity to one or more thermistors.

105. The automated medium-capacity assay processing unit of claims 103 or 104, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

106. The automated medium-capacity assay processing unit of claim 105, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

107. The automated medium-capacity assay processing unit of claims 104-106, wherein the one or more thermistors are two or more, or three or more thermistors.

108. The automated medium-capacity assay processing unit of claim 107, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

109. The automated medium-capacity assay processing unit of any of claims 103-108, wherein the one or more LEDs are present on a printer circuit board (PCB), wherein the PCB maintains the LEDs, the enclosed illumination chamber, and the consumable holder when the consumable holder is located at an imaging station at a third temperature.

110. The automated medium-capacity assay processing unit of claim 109, wherein the PCB comprises an aluminum core.1 11. The automated medium-capacity assay processing unit of any of claims 65-110, wherein the two or more imaging stations comprises a clamping component.

112. The automated medium-capacity assay processing unit of claim 111, wherein the clamping component engages with a consumable and provides a downward clamping force on the consumable when the consumable is present in the two or more imaging stations.

113. The automated medium-capacity assay processing unit of any of claims 101-112, wherein the secondary illumination component comprises one or more LEDs positioned beneath the consumable holder thereby enabling capture of a forward scatter image, wherein the consumable holder is in the third position114. The automated medium-capacity assay processing unit of any of claims 101-113, wherein the secondary illumination component comprises one or more LEDs positioned above the consumable holder thereby enabling capture of a back scatter image, wherein the consumable holder is in the third position1 15. The automated medium-capacity assay processing unit of claims 113 or 114, wherein the one or more LEDs are in proximity to one or more thermistors.

116. The automated medium-capacity assay processing unit of any of claims 113-115, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

117. The automated medium-capacity assay processing unit of claim 116, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

118. The automated medium-capacity assay processing unit of claims 116 or 117, wherein the one or more thermistors are two or more, or three or more thermistors.

119. The automated medium-capacity assay processing unit of claim 118, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

120. The automated medium-capacity assay processing unit of any of claims 113-119, wherein the one or more LEDs are present on a second printer circuit board (PCB).

121. The automated medium-capacity assay processing unit of claim 120, wherein the second PCB comprises an aluminum core.

122. The automated medium-capacity assay processing unit of any of claims 65-121, further comprising a degassing component.

123. The automated medium-capacity assay processing unit of any of claims 1-122, wherein the automated assay processing unit capable of being connected to one or more different automated assay processing units.

124. The automated medium-capacity assay processing unit of claim 123, wherein the one or more different automated medium-capacity assay processing units are adjacent to the automated medium-capacity assay processing unit such that the one or more automated medium-capacity assay processing units are parallel to the automated medium-capacity assay processing unit.

125. The automated medium-capacity assay processing unit of claim 123, wherein the one or more different automated medium-capacity assay processing units are in series to the automated medium-capacity assay processing unit such that the one or more automated mediumcapacity assay processing units are connected to the automated medium-capacity assay processing unit at an end.

126. The automated medium-capacity assay processing unit of claim 123, wherein the one or more different automated medium-capacity assay processing units are vertically offset from the automated medium-capacity assay processing unit such that the one or more automated medium-capacity assay processing units are stacked above or below the automated mediumcapacity assay processing unit.

127. The automated medium-capacity assay processing unit of any of claims 123-126, wherein the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.

128. The automated assay processing unit of any of claims 65-127, further comprising one or more consumables.

129. The automated assay processing unit of claim 128, wherein the one or more consumables are microfluidic devices.

130. An automated high-capacity assay processing unit, comprising: four or more fluid addition stations, two or more wash stations, wherein each wash station comprises a first carousel encircled by a second carousel, four or more imaging stations, and a consumable mover.

131. The automated high-capacity assay processing unit of claim 130, wherein the consumable mover comprises a conveyer system operably coupled to the fluid addition stations,wash stations, and imaging stations wherein the consumable mover is configured to move a consumable from the fluid addition station to the wash station and from the wash station to the imaging station.

132. The automated high-capacity assay processing unit of claim 131, wherein the conveyer system comprises a consumable holder that is capable of being moved between each station.

133. The automated high-capacity assay processing unit of claim 130, wherein the consumable mover comprises a griping component that is capable of gripping a consumable and moving the consumable between each station.

134. The automated high-capacity assay processing unit of any of claims 130-133, wherein the four or more fluid addition stations comprise one or more heating elements configured to heat one or more consumables to a first temperature, wherein the one or more consumables are present in the four or more fluid addition stations.

135. The automated high-capacity assay processing unit of any of claims 130-134, wherein the four or more fluid addition stations comprise one or more reagent dispensers comprising one or more reagent pumps fluidically connected to one or more reagent nozzles.

136. The automated high-capacity assay processing unit of claim 135, wherein the four or more fluid addition stations share the one or more reagent dispensers.

137. The automated high-capacity assay processing unit of claim 135, wherein each fluid addition station comprises the one or more reagent dispensers.

138. The automated high-capacity assay processing unit of any of claims 130-137, wherein the four or more fluid addition stations comprise a hydrophilic liquid dispenser comprising a hydrophilic liquid nozzle fluidically connected to a hydrophilic liquid pump.

139. The automated high-capacity assay processing unit of claim 138, wherein the hydrophilic liquid pump is configured to dispense a hydrophilic liquid through the hydrophilic liquid nozzle, wherein the consumable holder is located in a fluid addition station of the fluid addition stations.

140. The automated high-capacity assay processing unit of any of claims 130-139, wherein the four or more fluid addition stations comprise a mixing component.

141. The automated high-capacity assay processing unit of claim 140, wherein the four or more fluid addition stations share the mixing component.

142. The automated high-capacity assay processing unit of claim 140, wherein each fluid addition station comprises the mixing component.

143. The automated high-capacity assay processing unit of any of claims 140-142, wherein the mixing component comprises a mixing motor and an engagement arm that is configured to engage with a consumable, wherein the consumable is present in fluid addition station of the four or more fluid addition stations.

144. The automated high-capacity assay processing unit of claim 143, wherein the engagement arm engages with a portion of the consumable.

145. The automated high-capacity assay processing unit of claim 143, wherein the engagement arm engages with the entire consumable.

146. The automated high-capacity assay processing unit of claim 142, wherein the mixing component comprises a ballistic mixing device.

147. The automated high-capacity assay processing unit of claim 146, wherein the ballistic mixing device is a pipettor.

148. The automated high-capacity assay processing unit of any of claims 130-147, wherein the two or more wash stations comprise a first retractable magnet beneath a consumable, wherein the consumable is present in a wash station of the two or more wash stations, wherein the first retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state.

149. The automated high-capacity assay processing unit of claim 148, wherein the first retractable magnet is encased in a sleeve in the retracted state wherein the sleeve shields the first retractable magnet thereby reducing the local magnetic field.

150. The automated high-capacity assay processing unit of claim 148 or 149, wherein the two or more wash stations share the first retractable magnet.

151. The automated high-capacity assay processing unit of claim 148 or 149, wherein each wash station comprises the first retractable magnet.

152. The automated high-capacity assay processing unit of claim 148-151, further comprising a plurality of first retractable magnets.

153. The automated high-capacity assay processing unit of claims 130-151, wherein the two or more wash stations comprise one or more heating elements configured to heat one or more consumables to a second temperature, wherein the one or more consumables are present in the two or more wash stations.

154. The automated high-capacity assay processing unit of any of claims 130-153, wherein the two or more wash stations comprise a second retractable magnet.

155. The automated high-capacity assay processing unit of claim 154, wherein the second retractable magnet is positioned above a consumable when the consumable is in the two or more wash stations, wherein the second retractable magnet is configured to be proximate to the consumable in an un-retracted state and distal to the consumable in a retracted state.

156. The automated high-capacity assay processing unit of claim 154 or 155, wherein the second retractable magnet is positioned in a position that is horizontally offset from the first retractable magnet.

157. The automated high-capacity assay processing unit of any of claims 154-156, wherein the first retractable magnet and the second retractable magnet are configured to not be in the un-retracted state at the same time.

158. The automated high-capacity assay processing unit of any of claims 154-157, further comprising a plurality of second retractable magnets.

159. The automated high-capacity assay processing unit of any of claims 130-158, wherein two or more wash stations comprise an aspiration device comprising an aspiration nozzle fluidically connected to an aspiration pump.

160. The automated high-capacity assay processing unit of claims 159, wherein the aspiration pump is configured to aspirate a hydrophilic liquid through the aspiration nozzle, wherein a consumable is present in the two or more wash stations.

161. The automated high-capacity assay processing unit of claim 159 or 160, wherein the two or more wash stations share the aspiration device.

162. The automated high-capacity assay processing unit of claim 159 or 160, wherein each wash station comprises the aspiration device.

163. The automated high-capacity assay processing unit of any of claims 130-162, wherein the two or more wash stations comprise a hydrophobic liquid dispenser comprising a hydrophobic liquid nozzle fluidically connected to a hydrophobic liquid pump.

164. The automated high-capacity assay processing unit of claim 163, wherein the hydrophobic liquid pump is configured to dispense a hydrophobic liquid through the hydrophobic liquid nozzle, wherein a consumable is present in one of the two or more wash stations.

165. The automated high-capacity assay processing unit of claim 164, wherein the aspiration pump is configured to aspirate the hydrophilic liquid and the hydrophobic liquid pump is configured to dispense the hydrophobic liquid simultaneously, wherein a consumable is present in the two or more wash stations.

166. The automated high-capacity assay processing unit of any of claims 163-165, wherein the two or more wash stations share the hydrophobic liquid dispenser.

167. The automated high-capacity assay processing unit of any of claims 163-165, wherein each wash station comprises the hydrophobic liquid dispenser.

168. The automated high-capacity assay processing unit of any of claims 130-167, wherein the first carousel has a lower capacity for consumables than the second carousel.

169. The automated high-capacity assay processing unit of any of claims 130-168, wherein the first carousel rotates in lock-step with the second carousel.

170. The automated high-capacity assay processing unit of any of claims 130-169, wherein the first carousel rotates out of sync with the second carousel.

171. The automated high-capacity assay processing unit of any of claims 130-170, wherein the two or more imaging stations comprise an imaging component and a secondary illumination component.

172. The automated high-capacity assay processing unit of claim 171, wherein the imaging component is positioned beneath a consumable, wherein the consumable is present in one of the two or more imaging stations.

173. The automated high-capacity assay processing unit of claims 171 or 172, wherein the imaging component comprises an illumination component comprising one or more LEDs encased in an enclosed illumination chamber having an opening positioned beneath the third position.

174. The automated high-capacity assay processing unit of claim 173, wherein the one or more LEDs are in proximity to one or more thermistors.

175. The automated high-capacity assay processing unit of claims 173 or 174, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

176. The automated high-capacity assay processing unit of claim 175, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

177. The automated high-capacity assay processing unit of claims 174-176, wherein the one or more thermistors are two or more, or three or more thermistors.

178. The automated high-capacity assay processing unit of claim 177, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

179. The automated high-capacity assay processing unit of any of claims 173-178, wherein the one or more LEDs are present on a printer circuit board (PCB), wherein the PCB maintains the LEDs, the enclosed illumination chamber, and the consumable holder when the consumable holder is at the third position at a third temperature.

180. The automated high-capacity assay processing unit of claim 179, wherein the PCB comprises an aluminum core.

181. The automated high-capacity assay processing unit of any of claims 130-180, wherein the two or more imaging stations comprises a clamping component.

182. The automated high-capacity assay processing unit of claim 181, wherein the clamping component engages with a consumable and provides a downward clamping force on the consumable, wherein the consumable is present in the two or more imaging stations.

183. The automated high-capacity assay processing unit of any of claims 171-182, wherein the secondary illumination component comprises one or more LEDs positioned beneath the consumable holder thereby enabling capture of a forward scatter image, wherein the consumable holder is in the third position184. The automated high-capacity assay processing unit of any of claims 171-183, wherein the secondary illumination component comprises one or more LEDs positioned above the consumable holder thereby enabling capture of a back scatter image, wherein the consumable holder is in the third position185. The automated high-capacity assay processing unit of claims 183 or 184, wherein the one or more LEDs are in proximity to one or more thermistors.

186. The automated high-capacity assay processing unit of any of claims 183-185, wherein the one or more LEDs are two or more, three or more, four or more, five or more, or six or more LEDs.

187. The automated high-capacity assay processing unit of claim 186, wherein the two or more, three or more, four or more, five or more, or six or more LEDs are arranged in a circular pattern.

188. The automated high-capacity assay processing unit of claims 186 or 187, wherein the one or more thermistors are two or more, or three or more thermistors.

189. The automated high-capacity assay processing unit of claim 188, wherein the two or more or three or more thermistors are spaced apart such that the thermistors are between two LEDs.

190. The automated high-capacity assay processing unit of any of claims 184-189, wherein the one or more LEDs are present on a second printer circuit board (PCB).

191. The automated high-capacity assay processing unit of claim 190, wherein the second PCB comprises an aluminum core.

192. The automated high-capacity assay processing unit of any of claims 130-191, further comprising a degassing component.

193. The automated high-capacity assay processing unit of any of claims 1-192, wherein the automated assay processing unit capable of being connected to one or more different automated assay processing units.

194. The automated high-capacity assay processing unit of claim 193, wherein the one or more different automated high-capacity assay processing units are adjacent to the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are parallel to the automated high-capacity assay processing unit.

195. The automated high-capacity assay processing unit of claim 193, wherein the one or more different automated high-capacity assay processing units are in series to the automatedhigh-capacity assay processing unit such that the one or more automated high-capacity assay processing units are connected to the automated high-capacity assay processing unit at an end.

196. The automated high-capacity assay processing unit of claim 193, wherein the one or more different automated high-capacity assay processing units are vertically offset from the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are stacked above or below the automated high-capacity assay processing unit.

197. The automated high-capacity assay processing unit of any of claims 193-196, wherein the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.

198. The automated assay processing unit of any of claims 130-197, further comprising one or more consumables.

199. The automated assay processing unit of claim 198, wherein the one or more consumables are microfluidic devices.

200. The automated high-capacity assay processing unit of any of claims 130-199, further comprising a degassing component.

201. The automated high-capacity assay processing unit of any of claims 130-200, wherein the automated assay processing unit capable of being connected to one or more different automated assay processing units.

202. The automated high-capacity assay processing unit of claim 201, wherein the one or more different automated high-capacity assay processing units are adjacent to the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are parallel to the automated high-capacity assay processing unit.

203. The automated high-capacity assay processing unit of claim 202, wherein the one or more different automated high-capacity assay processing units are in series to the automatedhigh-capacity assay processing unit such that the one or more automated high-capacity assay processing units are connected to the automated high-capacity assay processing unit at an end.

204. The automated high-capacity assay processing unit of claim 203, wherein the one or more different automated high-capacity assay processing units are vertically offset from the automated high-capacity assay processing unit such that the one or more automated high-capacity assay processing units are stacked above or below the automated high-capacity assay processing unit.

205. The automated high-capacity assay processing unit of any of claims 201-204, wherein the one or more different automated assay processing units are at least two of: adjacent to the automated assay processing unit, in series to the automated assay processing unit, or vertically offset from the automated assay processing unit.

206. The automated assay processing unit of any of claims 130-205, further comprising one or more consumables.

207. The automated assay processing unit of claim 206, wherein the one or more consumables are microfluidic devices.

208. A method of measuring or detecting a target analyte in a sample, the method comprising: a) loading a consumable comprising wells into the automated assay processing unit of claims 1-64; b) dispensing the sample comprising the target analyte into a first region of the consumable, c) dispensing a particle solution comprising microparticles bound to a first specific binding partner into the first region, d) dispensing a conjugate solution comprising a second specific binding partner that is detectably labeled into a second region of the consumable, e) mixing and incubating the sample and the particle solution in the first region thereby binding the target analyte to the microparticles, f) subjecting the microparticles bound to the target analyte to a magnetic field to move the microparticles to the second region,g) mixing and incubating the microparticles bound to the target analyte with the conjugate solution thereby producing microparticles bound to detectably labeled target analyte, h) subjecting the microparticles bound to the detectably labeled target analyte to a magnetic field to move the microparticles to a third region of the consumable comprising wells wherein the wells comprise substrate solution that reacts with the detectably labeled target analyte to produce a detectable signal; i) sealing the wells using a hydrophobic liquid; and j) imaging the wells thereby detecting the detectable signal.

209. The method of claim 208, wherein the dispensing of step a) is performed by a sample dispenser.

210. The method of claim 208 or 209, wherein the dispensing of step b) is performed by a first reagent dispenser and the dispensing of step d) is performed by a second reagent dispenser.

211. The method of any of claims 208-210, wherein step d) further comprises dispensing the hydrophilic liquid into the third region and dispensing a wash buffer into at least a fourth region and a fifth region.

212. The method of any of claims 208-211, wherein the mixing of step e) comprises engaging the engagement arm of the mixing component with at least a portion of the consumable and vibrating or agitating the consumable.

213. The method of any of claims 208-211, wherein the mixing of step e) comprises contacting the first region with a ballistic mixing device of a mixing component and agitating the first region.

214. The method of any of claims 208-213, wherein the incubating of step e) comprises heating the consumable to a first temperature.

215. The method of claim 214, wherein the first temperature is 30 C-45 C.

216. The method of any of claims 208-215, wherein the subjecting the microparticles bound to the target analyte to a magnetic field step f) comprises positioning the first magnet underthe first region, un-retracting the first magnet, moving the first magnet or the consumable positioning the first magnet under the second region, and retracting the first magnet.

217. The method of any of claims 211-216, further comprising: k) subjecting the microparticles bound to the target analyte to a magnetic field to move the microparticles from the first region to the fourth region of the consumable: and l) mixing the microparticles bound to the target analyte with the wash buffer, wherein steps k) and 1) occur prior to step f).

218. The method of any of claims 208-217, wherein subjecting the microparticles bound to the detectably labeled target analyte to a magnetic field of step h) comprises positioning the first magnet under the second region, un-retracting the first magnet, moving the first magnet to the third region, and retracting the first magnet.

219. The method of any of claims 208-218, further comprising: m) subjecting the microparticles bound to the detectably labeled target analyte to a magnetic field to move the microparticles from the second region to the fifth region of the consumable; and n) mixing the microparticles bound to the target analyte with the wash buffer, wherein steps k) and 1) occur prior to step f).

220. The method of claim 219, wherein the mixing of step g), 1), and n) comprises positioning the second magnet over the second region, the fourth region, or the fifth region, un- retracting the second magnet, and retracting the second magnet.

221. The method of any of claims 208-220, wherein the incubating of step g) comprises heating the consumable to a second temperature.

222. The method of claim 221, wherein the second temperature is 30-45 C.

223. The method of any of claims 208-222, wherein the sealing the wells using a hydrophobic liquid of step i) comprises: i) dispensing from a hydrophobic liquid dispenser the hydrophobic liquid into the third region thereby flowing the hydrophobic liquid over the wells and displacing the hydrophilic liquid, andii) aspirating the hydrophilic liquid using an aspiration device.

224. The method of claim 223, wherein the dispensing of the hydrophobic liquid and the aspirating the hydrophilic liquid occurs simultaneously.

225. The method of any of claims 208-224, wherein imaging the wells of step j) comprises: i) immobilizing the consumable, ii) adjusting a focus of an imaging component to a first focal plane, iii) capturing a first scatter image, iv) capturing a first fluorescent image, and v) repeating steps ii-iv) one or more times to generate a stack of scatter images and fluorescent images spanning a plurality of focal planes.

226. The method of claim 225, wherein step iii) comprises enabling one or more scatter LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more scatter LEDs.

227. The method of claim 225 or 226, wherein step iv) comprises enabling one or more fluorescent LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more fluorescent LEDs.

228. The method of any of claims 225-227, wherein step i) comprises contacting an alignment datum with the consumable thereby activating a raising element and a clamping component, wherein: the raising element is positioned below the consumable and the clamping component is positioned above the consumable, and the raising element provides an upward force on the consumable and the clamping component provides a downward force on the consumable.

229. The method of any of claims 208-228, wherein steps a)-d) are performed in a first position on the automated assay processing unit.

230. The method of any of claims 208-229, wherein steps e)-i) are performed in a second position on the automated assay processing unit.

231. The method of any of claims 208-230, wherein step j) is performed in a third position on the automated assay processing unit.

232. The method of any of claims 208-231, further comprising moving the consumable from the first position to the second position following step d).

233. The method of any of claims 208-232, further comprising moving the consumable from the second position to the third position following step j).

234. The method of any of claims 208-233, wherein the particle solution further comprises assisting microparticles.

235. The method of any of claims 208-234, wherein the target analyte is a protein.

236. The method of any claims 208-235, wherein the first and second specific binding members are antibodies that specifically bind to different epitopes on the target analyte.

237. The method of any of claims 208-236, wherein the detectable label is an enzyme that reacts with the hydrophilic liquid.

238. The method of any of claims 208-237, wherein the hydrophobic liquid is oil.

239. The method of any of claims 208-238, wherein the hydrophilic liquid is a substrate solution.

240. The method of any of claims 208-239, wherein step b) further comprises dispensing lysis buffer into the first region of the consumable comprising the sample.

241. The method of any of claims 208-240, wherein step c) occurs before step b)242. The method of any of claims 208-240, wherein: the microparticles comprise a plurality of first microparticles and a plurality of second microparticles,the first specific binding partner of the plurality of first microparticles is specific to a first epitope and the first specific binding partner of the plurality of first microparticles is specific to a second epitope, and the plurality of first microparticles comprise a first fluorophore and the plurality of second microparticles comprise a second fluorophore.

243. The method of claim 242, wherein the first epitope and the second epitope are: the same epitope, or different epitopes.

244. The method of claim 243, wherein the target analyte is a first target analyte and a second target analyte and the first target analyte is different from the second target analyte.

245. The method of any of claims 242-244, wherein the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles.

246. The method of any of claims 242-245, wherein the plurality of first microparticles are coated with the first fluorophore and the plurality of second microparticles are coated with the second fluorophore.

247. The method of any of claims 242-245, wherein the plurality of first microparticles are embedded with the first fluorophore and the plurality of second microparticles are embedded with the second fluorophore.

248. The method of any of claims 242-247, wherein capturing the first fluorescent image of step j) further comprises capturing a second fluorescent and capturing a third fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, and the third fluorescent image detects the plurality of second microparticles.

249. The method of claim 248, wherein capturing the first scatter image of step j) further comprises capturing a second scatter image and capturing a third scatter image; wherein the second scatter image is taken after the first fluorescent image, the third scatter image is taken after the first fluorescent image.

250. The method of any of claims 208-240, wherein: the microparticles comprise a plurality of first microparticles, a plurality of second microparticles, and a plurality of third microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first epitope, the first specific binding partner of the plurality of second microparticles is specific to a second epitope, and the first specific binding partner of the plurality of third microparticles is specific to a third epitope and the plurality of first microparticles comprise a first fluorophore, the plurality of second microparticles comprise a second fluorophore, and the plurality of third microparticles comprise a third fluorophore .

251. The method of claim 250, wherein the first epitope, the second epitope, the third epitope are the same epitope.

252. The method of claim 251, wherein the first epitope, the second epitope, and the third epitope are different epitopes.

253. The method of claim 252, wherein the target analyte is a first target analyte, a second target analyte, and a third target analyte and the first target analyte, the second target analyte and the third target analyte are different.

254. The method of any of claims 250-253, wherein the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles and the plurality of third microparticles.

255. The method of any of claims 250-254, wherein the plurality of first microparticles are coated with the first fluorophore, the plurality of second microparticles are coated with the second fluorophore, and the plurality of third microparticles are coated with the third fluorophore.

256. The method of any of claims 250-255, wherein the plurality of first microparticles are embedded with the first fluorophore, the plurality of second microparticles are embedded with the second fluorophore, and the plurality of third microparticles are embedded with the third fluorophore.

257. The method of claims 250-256, wherein the second fluorophore and the third fluorophore produce the same emission wavelength but are excited by different excitation wavelengths.

258. The method of any of claims 242-247, wherein capturing the first fluorescent image of step j) further comprises capturing a second fluorescent, capturing a third fluorescent image, and capturing a fourth fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, the third fluorescent image detects the plurality of second microparticles, and the fourth florescent image detects the plurality of third microparticles.

259. The method of claim 258, wherein capturing the first scatter image of step j) further comprises capturing a second scatter image, capturing a third scatter image, and capturing a fourth scatter image; wherein the second scatter image is taken after the first fluorescent image, the third scatter image is taken after the second fluorescent image, and the fourth scatter image is taken after the third fluorescent image.

260. A method of measuring or detecting a target analyte in a sample, the method comprising: a) loading a consumable comprising wells into the automated assay processing unit of claims 1-64, b) dispensing the sample comprising the target analyte into a first region of the consumable, c) dispensing a particle solution comprising microparticles bound to a first specific binding partner into the first region, d) mixing and incubating the sample and the particle solution in the first region thereby hybridizing the target analyte to the microparticles, e) subjecting the microparticles bound to the target analyte to a magnetic field to move the microparticles to a second region of the consumable comprising wells, f) dispensing reagents into the third region that amplify and detect the target analyte under isothermal amplification conditions comprising a detection probe that produces a detectable signal following amplification of the target analyte, g) sealing the wells using a hydrophobic liquid; and h) imaging the wells thereby detecting the detectable signal.

261. The method of claim 260, wherein the target analyte is a nucleic acid.

262. The method of claim 261 , wherein the nucleic acid is DNA or RNA.

263. The method of any of claims 260-262, wherein the first specific binding member is a nucleic acid that specifically hybridizes to the target analyte.

264. The method of any of claims 260-263, wherein the dispensing of step a) is performed by a sample dispenser.

265. The method of any of claims 260-264, wherein the dispensing of step b) is performed by a first reagent dispenser.

266. The method of any of claims 260-265, wherein step d) further comprises and dispensing a wash buffer into at least a third region.

267. The method of any of claims 260-266, wherein the mixing of step d) comprises engaging the engagement arm of a mixing component with at least a portion of the consumable and vibrating or agitating the consumable.

268. The method of any of claims 260-266, wherein the mixing of step d) comprises contacting the first region with a ballistic mixing device of a mixing component and agitating the first region.

269. The method of any of claims 260-268, wherein the incubating of step d) comprises heating the consumable to a first temperature.

270. The method of claim 269, wherein the first temperature is xx.

271. The method of any of claims 260-270, wherein the subjecting the microparticles bound to the target analyte to a magnetic field step f) comprises positioning the first magnet under the first region, un-retracting the first magnet, moving the first magnet or the consumable positioning the first magnet under the second region, and retracting the first magnet.

272. The method of any of claims 260-271, further comprising:i) subjecting the microparticles bound to the target analyte to a magnetic field to move the microparticles from the first region to the third region of the consumable; and j) mixing the microparticles bound to the target analyte with the wash buffer, wherein steps i) and j) occur prior to step e).

273. The method of any of claims 260-272, wherein subjecting the microparticles bound to the detectably labeled target analyte to a magnetic field of step h) comprises positioning the first magnet under the second region, un-retracting the first magnet, moving the first magnet to the third region, and retracting the first magnet.

274. The method of claim 272 or 273, wherein the mixing of step j) comprises positioning the second magnet over the fourth region, un-retracting the second magnet, and retracting the second magnet.

275. The method of any of claims 272-274, wherein the incubating of step j) comprises heating the consumable to a second temperature.

276. The method of claim 275, wherein the second temperature is xx.

277. The method of any of claims 272-276, wherein the dispensing of step f) comprises: k) dispensing from one or more reagent dispensers the reagents into the second region thereby flowing the reagents into the wells.

278. The method of any of claims 272-277, wherein the isothermal amplification is recombinase polymerase amplification.

279. The method of claim 278, wherein the reagents comprise dNTPs, a recombinase, a single-stranded DNA binding protein, a strand-displacing polymerase, a forward and reverse primer specific to the target analyte, and a detection probe that produces a detectable signal following amplification of the target analyte.

280. The method of any of claims 272-278, wherein the isothermal amplification is loop-mediated isothermal amplification.

281. The method of claim 280, wherein the reagents comprise dNTPs, at least four primers that recognize at least 6 distinct regions in the target analyte, a strand-displacing polymerase, and a detection probe that produces a detectable signal following amplification of the target analyte.

282. The method of any of claims 272-278, wherein the isothermal amplification is helicase dependent amplification.

283. The method of claim 282, wherein the reagents comprise dNTPs, one or more helicases, a forward and reverse primer specific to the target analyte, a polymerase, and a detection probe that produces a detectable signal following amplification of the target analyte.

284. The method of any of claims 272-278, wherein the isothermal amplification is nicking enzyme amplification reaction.

285. The method of claim 284, wherein the reagents comprise dNTPs, a DNA polymerase, a first amplification oligonucleotide comprising a first nicking enzyme site, a second amplification oligonucleotide comprising a second nicking enzyme site, a first nicking enzyme that recognizes the first nicking enzyme site and second nicking enzyme recognizing the second enzyme site, and a detection probe that produces a detectable signal following amplification of the target analyte.

286. The method of any of claims 272-285, wherein the sealing the wells using a hydrophobic liquid of step g) comprises:1) dispensing from a hydrophobic liquid dispenser the hydrophobic liquid into the second region thereby flowing the hydrophobic liquid over the wells and displacing the reagents.

287. The method of any of claims 272-286, wherein imaging the wells of step h) comprises: i) immobilizing the consumable, ii) adjusting a focus of an imaging component to a first focal plane, iii) capturing a first scatter image, iv) capturing a first fluorescent image, and v) repeating steps ii-iv) one or more times to generate a stack of scatter images and fluorescent images spanning a plurality of focal planes.

288. The method of claim 287, wherein step iii) comprises enabling one or more scatter LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more scatter LEDs.

289. The method of claim 287 or 288, wherein step iv) comprises enabling one or more fluorescent LEDs present in the imaging component, capturing the first scatter image, and disabling the one or more fluorescent LEDs.

290. The method of any of claims 287-289, wherein step i) comprises contacting an alignment datum with the consumable thereby activating a raising element and a clamping component, wherein: the raising element is positioned below the consumable and the clamping component is positioned above the consumable, and the raising element provides an upward force on the consumable and the clamping component provides a downward force on the consumable.

291. The method of any of claims 272-290, wherein steps a)-c) are performed in a first position on the automated assay processing unit.

292. The method of any of claims 272-291, wherein steps d)-g) are performed in a second position on the automated assay processing unit.

293. The method of any of claims 272-292, wherein step h) is performed in a third position on the automated assay processing unit.

294. The method of any of claims 272-293, further comprising moving the consumable from the first position to the second position following step c).

295. The method of any of claims 272-294, further comprising moving the consumable from the second position to the third position following step g).

296. The method of any of claims 272-295, wherein the particle solution further comprises assisting microparticles.

297. The method of any of claims 272-296, wherein the hydrophobic liquid is oil.

298. The method of any of claims 272-297, wherein step b) further comprises dispensing lysis buffer into the first region of the consumable comprising the sample.

299. The method of any of claims 272-298, wherein: the microparticles comprise a plurality of first microparticles and a plurality of second microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first nucleic acid sequence and the first specific binding partner of the plurality of first microparticles is specific to a nucleic acid sequence, and the plurality of first microparticles comprise a first fluorophore and the plurality of second microparticles comprise a second fluorophore.

300. The method of claim 299, wherein the first nucleic acid sequence and the second nucleic acid sequence are the same nucleic acid sequence.

301. The method of claim 300, wherein the first nucleic acid sequence and the second nucleic acid sequence are different nucleic acid sequences.

302. The method of claim 301, wherein the target analyte is a first target analyte and a second target analyte and the first target analyte is different from the second target analyte.

303. The method of any of claims 299-302, wherein the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles.

304. The method of any of claims 299-303, wherein the plurality of first microparticles are coated with the first fluorophore and the plurality of second microparticles are coated with the second fluorophore.

305. The method of any of claims 299-304, wherein the plurality of first microparticles are embedded with the first fluorophore and the plurality of second microparticles are embedded with the second fluorophore.

306. The method of any of claims 299-305, wherein capturing the first fluorescent image of step j) further comprises capturing a second fluorescent and capturing a third fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, and the third fluorescent image detects the plurality of second microparticles.

307. The method of claim 306, wherein capturing the first scatter image of step j) further comprises capturing a second scatter image and capturing a third scatter image; wherein the second scatter image is taken after the first fluorescent image, the third scatter image is taken after the first fluorescent image.

308. The method of any of claims 260-307, wherein step c) occurs before step b).

309. The method of any of claims 272-298, wherein: the microparticles comprise a plurality of first microparticles, a plurality of second microparticles, and a plurality of third microparticles, the first specific binding partner of the plurality of first microparticles is specific to a first nucleic acid sequence, the first specific binding partner of the plurality of second microparticles is specific to a second nucleic acid sequence, and the first specific binding partner of the plurality of third microparticles is specific to a third nucleic acid sequence and the plurality of first microparticles comprise a first fluorophore, the plurality of second microparticles comprise a second fluorophore, and the plurality of third microparticles comprise a third fluorophore .

310. The method of claim 309, wherein the first nucleic acid sequence, the second nucleic acid sequence, the third nucleic acid sequence are the same nucleic acid sequence.

311. The method of claim 310, wherein the first nucleic acid sequence, the second nucleic acid sequence, and the third nucleic acid sequence are different nucleic acid sequences.

312. The method of claim 311, wherein the target analyte is a first target analyte, a second target analyte, and a third target analyte and the first target analyte, the second target analyte and the third target analyte are different.

313. The method of any of claims 309-312, wherein the plurality of first microparticles have a different abundance of first specific binding partners than the plurality of second microparticles and the plurality of third microparticles.

314. The method of any of claims 309-313, wherein the plurality of first microparticles are coated with the first fluorophore, the plurality of second microparticles are coated with the second fluorophore, and the plurality of third microparticles are coated with the third fluorophore.

315. The method of any of claims 309-314, wherein the plurality of first microparticles are embedded with the first fluorophore, the plurality of second microparticles are embedded with the second fluorophore, and the plurality of third microparticles are embedded with the third fluorophore.

316. The method of claims 309-315, wherein the second fluorophore and the third fluorophore produce the same emission wavelength but are excited by different excitation wavelengths.

317. The method of any of claims 309-316, wherein capturing the first fluorescent image of step j) further comprises capturing a second fluorescent, capturing a third fluorescent image, and capturing a fourth fluorescent image; wherein the first fluorescent image detects the detectably labeled target analyte, the second fluorescent image detects the plurality of first microparticles, the third fluorescent image detects the plurality of second microparticles, and the fourth florescent image detects the plurality of third microparticles.

318. The method of claim 317, wherein capturing the first scatter image of step j) further comprises capturing a second scatter image, capturing a third scatter image, and capturing a fourth scatter image; wherein the second scatter image is taken after the first fluorescent image, the third scatter image is taken after the second fluorescent image, and the fourth scatter image is taken after the third fluorescent image.

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