Systems and methods of sample depositing and testing

Abstract

The present application is generally directed to systems, methods, and devices for diagnostics for sensing and / or identifying pathogens, genomic materials, proteins, and / or other small molecules or biomarkers, which may utilize loop-mediated isothermal amplification (LAMP). In some implementations, additional components are added to the sample, or reagents, which aid in the diagnostic procedures, mixing, sample deposition, or compensation for inhibitors that may be present. Also disclosed herein are nucleic acid primers, which may be provided in the devices and, which sensitively and specifically detect pathogens in biological samples by LAMP amplification. The biological samples may be derived from subjects including humans, plants, food, soil, contaminated surfaces, or animals including livestock.

Description

[0001] SYSTEMS AND METHODS OF SAMPLE DEPOSITING AND TESTING

[0002] FIELD

[0003]

[0001] The present application is generally directed to systems, methods, and devices for sensing and / or identifying pathogens, genomic materials, proteins, and / or other small molecules or biomarkers and using historical sensing and testing results to track a subject’s or product’s wellness score or likelihood of being infected with a trackable pathogen. The present application also relates to sample preparation.

[0004] BACKGROUND

[0005] [2] Pathogens in a sample may be identified by detecting specific genomic material (DNA or RNA). In conventional nucleic acid testing (“NAAT”), genomic material in a sample may first be exponentially copied using a molecular amplification process known as the polymerase chain reaction (“PCR”) until the quantity of DNA present is great enough to be measurable. In the case of RNA, the genomic material of many viruses, an additional step can be included to first transcribe the RNA into DNA before amplifying by PCR. As an alternative, loop-mediated isothermal amplification (LAMP) offers several advantages over PCR for pathogen detection purposes, including the ability to perform the amplification reaction at a non-cyclical and relatively low temperature. There is a lasting need for improved pathogen detection methods and tools, e.g., using LAMP, particularly in geographical regions where the use of complex laboratory equipment is not feasible.

[0006] [3] Monitoring of viral avian flu in poultry populations has been of increased recent interest due to numerous outbreaks of the disease.

[0007] SUMMARY

[0008] [4] According to one aspect of the disclosure, a method for identifying a target molecule in a biological sample is disclosed. The method can include: filtering the biological sample to remove at least a portion of solid particles, the solid particles present in the sample; introducing a buffer to the sample to create a first mixture, the buffer including at least octylphenoxy poly(ethyleneoxy)ethanol; depositing the first mixture into a sample receptacle of a disposable cartridge; inserting the disposable cartridge into a cartridge receptacle of an analyzer device; mixing the first mixture with dried reagents including bovine serum albumin (BSA) and thereby generating a second mixture including the first mixture and the BSA; conveying at least a portion of the second mixture to at least one testing well, which includes at least one dried enzyme and / or a detection agent, such as one or more primers, an antibody or a binding fragment of an antibody, e.g., an ScFv, to generate a third mixture; optionally, increasing a temperature of the at least one testing well to a level sufficient for primer extension or amplification; and measuring an electrical characteristic of at least the portion of the third mixture in the at least one testing well, preferably by measuring a change in impedance at the at least one testing well.

[0009] [5] In some embodiments, filtering can include using a device, which is not integrated with the disposable cartridge. In further embodiments, the filtering can include using a column filter. In further embodiments, the filtering can include propelling the biological sample through a filter of a vial adaptor, the vial adaptor including: a cap member configured to engage the disposable cartridge; and a piston including a distal end, a proximal end, a body extending between the distal end and the proximal end, a channel, and a filter positioned in the channel, where the channel can allow flow of fluid through the piston and between a distal aperture formed on the distal end of the piston and a proximal aperture formed on the proximal end of the piston, where the piston can translate distally through the cap member and towards the vial, and where the filter can block passage of solid particles as the fluid flows through the piston; where a distal translation of the piston can cause a positive pressure change in the vial and withdrawal of fluid stored in the vial through the channel of the piston. In further embodiments, the filtering includes pipetting the sample through a filter of a filter adapter that can engage the disposable cartridge and a pipette.

[0010] [6] In some embodiments, the filtering is performed using a filter, which is internally disposed within the disposable cartridge. In further embodiments, the filter includes a sintered porous plug having plug pore sizes of 20-50 pm in diameter, such as 20, 30, 40, or 50 pm in diameter or having a pore size diameter that is within a range of pore size diameters defined by any two of the aforementioned pore size diameters.

[0011] [7] In some embodiments, the method includes rupturing a reagent blister to introduce the buffer to the sample and / or to rehydrate the dried reagents. In further embodiments, insertion of the disposable cartridge into the cartridge receptacle causes the rupturing of the reagent blister, the generating of the second mixture, and the conveying of at least the portion of the second mixture to the at least one testing well to generate the third mixture. [8] In some embodiments, the buffer includes 0.1% to 5% of the octylphenoxy poly(ethyleneoxy)ethanol by volume. In further embodiments, the buffer includes 1% of octylphenoxy poly(ethyleneoxy)ethanol by volume. In some embodiments, the dried reagents include 0.15% to 0.45% BSA by weight. In further embodiments, the dried reagents include 0.30% BSA by weight. In some embodiments, the biological sample is diluted prior to introduction into the sample receptacle. In further embodiments, the biological sample is diluted in water.

[0012] [9] In some embodiments, the biological sample includes solid particles, which are larger than 50 pm in diameter. In some embodiments, the buffer is dried and is rehydrated prior to or concomitantly with generation of the first mixture. In some embodiments, the biological sample is a cloacal sample. In further embodiments, the biological sample is an avian cloacal sample. In yet further embodiments, the biological sample is a chicken cloacal sample.

[0013]

[0010] According to another aspect of the present disclosure, an assay cartridge for analyzing a biological sample including a target nucleic acid is provided. The assay cartridge can include: a cartridge body that can be received by a reader device, the cartridge body including: at least one test well including an excitation electrode and a sensing electrode, where the at least one test well can receive at least a portion of the biological sample including the target nucleic acid; a sample introduction area that can receive a carrier containing the biological sample; and a fluid path fluidically coupling the sample introduction area to the test well, where the fluid path includes a filter that can inhibit solid particles of the sample from flowing to the test well; at least one reagent receptacle including at least one dried reagent, where the at least one dried reagent includes bovine serum albumin (BSA); and a buffer blister configured to be ruptured when the cartridge body is inserted into the reader device, the buffer blister including at least octylphenoxy poly(ethyleneoxy)ethanol, and where the rupturing of the reagent blister produces a force that mixes the at least one dried reagent with the buffer and with the biological sample to generate a mixture and moves at least a portion of the mixture through the fluid path to the at least one test well. In some embodiments, the assay cartridge can introduce the buffer to the sample.

[0011] According to another aspect of the present disclosure, an assay cartridge for analyzing a biological sample, the biological sample including a target nucleic acid, is provided, the assay cartridge including: a cartridge body that can be received by a reader device, the cartridge body including: at least one test well including an excitation electrode and a sensing electrode, where the at least one test well can receive at least a portion of the biological sample including the target nucleic acid, where the biological sample includes at least octylphenoxy poly(ethyleneoxy)ethanol; a sample introduction area that can receive a carrier containing the biological sample; and a fluid path fluidically coupling the sample introduction area to the test well, where the fluid path includes a filter configured to inhibit solid particles of the sample from flowing to the test well; at least one reagent receptacle including at least one dried reagent, where the at least one dried reagent includes bovine serum albumin (BSA); and a buffer blister configured to be ruptured when the cartridge body is inserted into the reader device, the buffer blister including a buffer, and where the rupturing of the reagent blister produces a force that mixes the at least one dried reagent with the buffer and with the biological sample to generate a mixture and moves at least a portion of the mixture through the fluid path to the at least one test well.

[0014]

[0012] In embodiments of an assay cartridge in accordance with the present disclosure, the filter can include plug pore sizes of 50 pm in diameter or smaller, such as less than 50, 40, 30, or 20 pm in diameter. In some embodiments, the filter includes plug pore sizes of 20 pm in diameter or larger, such as larger than 20, 30, 40, or 50 pm in diameter. In some embodiments, the biological sample includes solid particles, preferably above 50 pm in diameter. In some embodiments, the at least one dried reagent includes 0.15% to 0.45% BSA by weight. In further embodiments, the at least one dried reagent includes 0.30% BSA by weight. In some embodiments, the biological sample is a cloacal sample. In further embodiments, the biological sample is an avian cloacal sample. In further embodiments, the biological sample is a chicken cloacal sample.

[0015]

[0013] In some embodiments, an assay cartridge of the present disclosure can include: a channel; and a cap configured to hold the carrier containing the biological sample, the cap further configured to mechanically couple to the cartridge body, where mechanically coupling the cap to the cartridge body causes compression of a trapped volume of a fluid to drive at least a portion of the sample through the fluid path into the test well, and where the cap includes: a retaining well having an interior diameter larger than an exterior diameter of the channel; and a retaining structure disposed within the retaining well and configured to retain the channel at a position spaced from a side interior wall and a rear interior wall of the retaining well to form at least one air channel fluidically coupled to an inner end of the channel.

[0016]

[0014] In some embodiments, the cartridge body includes a base and a translucent cover, the translucent cover including a planar surface defining one side of at least one of the test well and the fluid path. In some embodiments, the cartridge body includes at least a second test well containing an excitation electrode and a sensing electrode, and a second fluid path fluidically coupling the sample introduction area to the second test well, where the second test well can contain at least a portion of the sample, including the target nucleic acid. In some embodiments, the cartridge body includes a base and a printed circuit board (PCB), the PCB including a planar surface defining one side of at least one of the test well and the fluid path. In further embodiments, the PCB includes a heating element configured to heat the test well. In further embodiments, the PCB includes the excitation electrode and the sensing electrode. In further embodiments, the test well can mix a reagent and the sample into a substantially evenly mixed test fluid. In yet further embodiments, the reagent includes one or more dried reagents stored within the test well. In yet further embodiments, the cartridge body includes a plurality of test wells, and where at least a first test well of the plurality of test wells stores a reagent different from a reagent stored in a second test well of the plurality of test wells. In further embodiments, the cartridge body includes a plurality of test wells, and where at least two test wells of the plurality of test wells store the same reagent.

[0017]

[0015] In some embodiments, the cartridge body further includes a mixing chamber positioned between the sample introduction area and the test well along the fluid path, the mixing chamber configured to mix a reagent and the biological sample into a substantially evenly mixed test fluid. In a further embodiment, the reagent includes one or more dried reagents stored within the mixing chamber. In some embodiments, the assay cartridge includes a first electrode interface including a first contact pad leading to the excitation electrode and a second contact pad leading to the sensing electrode. In some embodiments, the assay cartridge further includes a gas-permeable, liquid-impermeable vent fluidically coupled to the test well. In some embodiments, the assay cartridge further includes a machine-readable cartridge identifier printed thereon, the cartridge identifier associated with one or more test protocols. In some embodiments, the assay cartridge is a disposable singleuse assay cartridge.

[0018]

[0016] According to another aspect of the present disclosure, a buffer vial configured to filter a biological sample prior to transferring the biological sample to a receiving device is disclosed. The buffer vial can include: a vial; a cap member that can removably couple with an opening of the vial; and a filter coupled to a rod, the rod slidingly coupled to the cap, where the rod can, when the cap member is coupled to the vial, push the filter from a first position by an open end of the vial to a second position closer to a closed end of the vial than the first position, where, at the second position, the filter confines solid particles of the sample to a portion of the vial proximate the closed end.

[0019]

[0017] In some embodiments, the filter includes plug pore sizes of 50 pm or smaller, such as less than 50, 40, 30, or 20 pm in diameter. In some embodiments, the filter includes plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter. In some embodiments, the rod can decouple from the cap member when the filter is in the second position. In some embodiments, the buffer vial further includes a filter support configured to inhibit curvature of the filter as the filter is pushed by the rod. In some embodiments, the biological sample is a cloacal sample. In further embodiments, the biological sample is an avian cloacal sample. In yet further embodiments, the biological sample is a chicken cloacal sample. In some embodiments, the vial includes a non-denaturing detergent. In some embodiments, the non- denaturing detergent is octylphenoxy poly(ethyleneoxy)ethanol.

[0020]

[0018] According to another aspect of the present disclosure, a vial adapter that can filter a biological sample, and transfer the biological sample, from a vial to a receiving device is provided. The vial adapter includes: a cap member configured to removably couple with an opening of a vial; and a piston including a distal end, a proximal end, a body extending between the distal end and the proximal end, a channel, and a filter positioned in the channel, the channel configured to allow flow of fluid through the piston and between a distal aperture formed on the distal end of the piston and a proximal aperture formed on the proximal end of the piston, the piston configured to translate distally through the cap member and towards the vial, and the filter configured to block passage of solid particles as the fluid flows through the piston; where a distal translation of the piston can cause a positive pressure change in the vial and withdrawal of fluid stored in the vial through the channel of the piston.

[0021]

[0019] In some embodiments, the filter includes plug pore sizes of 50 pm or smaller such as less than 50, 40, 30, or 20 pm in diameter. In some embodiments, the filter includes plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter. In some embodiments, the cap member includes an arm, where the piston includes a rack, and where the arm can engage the rack to allow the piston to translate distally through the cap member towards the vial and prevent the piston to translate proximally through the cap member away from the vial. In some further embodiments, the arm includes a tip, and where the tip is curved inwards towards an axis parallel to the body of the piston. In some further embodiments, the rack includes a plurality of teeth, where each of the plurality of teeth includes a leading edge and a stopping edge, and where the arm can: slide along the leading edges of the plurality of teeth and the piston is translated distally; and abut one of the stopping edges of the plurality of teeth to prevent the piston from translating proximally.

[0022]

[0020] In some embodiments, the proximal end of the piston can removably couple with a receiving device. In some embodiments, the piston includes a stopper extending radially and circumferentially outwards from the body of the piston, where the stopper can abut the cap member when the piston is in its distal-most position with respect to the cap member. In some embodiments, a width of the channel increases from the distal aperture to the proximal aperture. In some embodiments, the vial adaptor further includes a cover, where the cover can cover the distal end of the body of the piston. In further embodiments, the cover includes or is a molded elastomer.

[0023]

[0021] In some embodiments, the cover can abut an inner surface of the vial to create a seal between the cover and the inner surface of the vial. In some embodiments, the biological sample is a cloacal sample. In further embodiments, the biological sample is an avian cloacal sample. In yet further embodiments, the biological sample is a chicken cloacal sample.

[0024]

[0022] According to another aspect of the present disclosure, a filter adapter for filtering a biological sample having solid particles is provided. The filter adapter can include: a first portion that can receive a sample collection device; a second portion adapted for a cartridge sample port; and a filter positioned in a fluid path of the filter adapter, the filter being configured to separate solid particles from a biological sample fluid.

[0025]

[0023] In some embodiments, the filter includes plug pore sizes of 50 pm or smaller such as less than 50, 40, 30, 20, or 10 pm in diameter. In some embodiments, the filter includes plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter.

[0026]

[0024] In some embodiments, the biological sample is a cloacal sample. In some further embodiments, the biological sample is an avian cloacal sample. In yet further embodiments, the biological sample is a chicken cloacal sample.

[0027]

[0025] In some embodiments, the filter adapter further includes a filter support positioned within the fluid path and downstream the filter, where an upstream surface of the filter support contacts the filter. In some embodiments, the first portion and second portion are configured to interface each other via a threaded connection. In some embodiments, the first portion can create a fluidic seal with a first end of a shell of the filter, and the second portion can create a fluidic seal with a second end of a shell of the filter. In some embodiments, the sample collection device is a pipette. In other embodiments, the sample collection device is a syringe. In other embodiments, the sample collection device is an assay cartridge in accordance with the present disclosure.

[0028] BRIEF DESCRIPTION OF THE DRAWINGS

[0029]

[0026] FIGs. 1A-1E depict an example of another type or format of cartridge configured to detect a target that can be used in conjunction with a handheld system disclosed herein.

[0030]

[0027] FIGs. 1 F-l J schematically depict an example process of collecting and testing a sample.

[0031]

[0028] FIGs. 1K-1L depict an example cartridge with an example retainer that can hold a swab in place inside a swab receptacle.

[0032]

[0029] FIGs. 2A-2D depict example cartridges including internal filters for filtering sample fluid.

[0033]

[0030] FIGs. 3A and 3B depict an example of another handheld system disclosed herein.

[0031] FIGs. 4A-4N depict various examples of electrodes that can be used in a test well or channel of another suitable target detection cartridge as described herein.

[0034]

[0032] FIG. 5A depicts a first electrode or excitation electrode and a second electrode or signal electrode that may be spaced apart from one another within a test well or channel of another suitable target detection cartridge as described herein.

[0035]

[0033] FIG. 5B depicts an example signal that can be extracted from the signal electrode of FIG. 5 A.

[0036]

[0034] FIG. 5C depicts the resistance and reactance components extracted from a signal as shown in FIG 5B generated based on an example positive test.

[0037]

[0035] FIG. 5D depicts the resistance and reactance components extracted from signals as shown in FIG 5B from example tests of positive and negative controls.

[0038]

[0036] FIG. 5E depicts the resistance and reactance components extracted from a signal as shown in FIG 5B generated based on another example positive test.

[0039]

[0037] FIG. 6 depicts a schematic block diagram of an example reader device that can be used with the cartridges described herein.

[0040]

[0038] FIG. 7A depicts a flowchart of an example process for operating a reader device during a test as described herein.

[0041]

[0039] FIG. 7B depicts a flowchart of an example process for analyzing test data to detect a target as described herein.

[0042]

[0040] FIG. 8 depicts an example network diagram of a networked system for tracking infection potential.

[0043]

[0041] FIG. 9 is a flow diagram showing example interactions between components of the networked system of FIG. 8.

[0044]

[0042] FIG. 10 depicts a general architecture of a computing device implementing one or more of the components of the system of FIG. 8.

[0045]

[0043] FIGs. 11A-11F are flow diagrams representing example processes of collecting and testing a sample.

[0046]

[0044] FIG. 12 illustrates an example swab to be used with cartridges and methods of the present disclosure.

[0047]

[0045] FIGs. 13A and 13B illustrates an example buffer vial for filtering samples.

[0046] FIGs. 14A and 14B illustrate various views of an example vial adapter including a rotating piston coupled to a vial.

[0048]

[0047] FIG. 15 illustrates an exploded view of the vial adapter of FIGs. 14A and 14B, showing various components of the vial adapter.

[0049]

[0048] FIG. 16 illustrates a perspective view of the vial adapter of FIGs. 14A and 14B.

[0050]

[0049] FIG. 17A illustrates a perspective view of a piston portion of the vial adapter of FIGs. 14A and 14B.

[0051]

[0050] FIG. 17B illustrates the piston portion of FIG. 17A showing a seal separated from a piston body.

[0052]

[0051] FIG. 18A illustrates a cross-sectional view of the syringe portion of FIGs. 17A and 17B along the lines 18A-18A.

[0053]

[0052] FIG. 18B illustrates a cross-sectional view of the syringe portion of FIG. 17B along the lines 18B-18B.

[0054]

[0053] FIGs. 19A and 19B illustrate various views of a cap portion of the vial adapter of FIGs. 14A and 14B.

[0055]

[0054] FIG. 20A illustrates a cross-sectional view of the vial adapter of FIGs. 14A and 14B prior to actuation of a piston of the vial adapter.

[0056]

[0055] FIG. 20B illustrates a cross-sectional view of the vial adapter of FIGs. 14A and 14B during actuation of a piston of the vial adapter.

[0057]

[0056] FIG. 20C illustrates a cross-sectional view of the vial adapter of FIGs. 14A and 14B showing a piston of the vial adapter in its distal-most position.

[0058]

[0057] FIG. 21A illustrates the vial adapter of FIGs. 14A and 14B and a receiving device.

[0059]

[0058] FIG. 21B illustrates the vial adapter of FIGs. 21A and 21B coupled to a receiving device.

[0060]

[0059] FIG. 22 illustrates an example method of transferring fluid from a vial to a receiving device using the vial adapter of FIGs. 14A and 14B.

[0061]

[0060] FIGs. 23A-23C illustrate an example filter adaptor.

[0062]

[0061] FIGs. 24A and 24B illustrate another example filter adaptor.

[0062] FIG. 25 illustrates a time to reaction of an assay run with varying concentrations of bovine serum albumin.

[0063]

[0063] FIG. 26 illustrates time to reaction of an assay run with varying concentrations of octylphenoxy poly(ethyleneoxy)ethanol.

[0064]

[0064] FIG. 27 illustrates time to reaction of different assays run at varying combinations of bovine serum albumin concentration and octylphenoxy poly(ethyleneoxy)ethanol concentration.

[0065]

[0065] FIGs. 28A and 28B illustrates time to reaction of an assay run using varying elution buffer volumes.

[0066]

[0066] FIG. 29 illustrates time to reaction an assay where the sample was processed using a gel column.

[0067] DETAILED DESCRIPTION

[0068]

[0067] Aspects of the disclosure herein concern the use of amplification and contactless electrical sensing to detect the presence and / or amount of a target in a sample. More specifically, the systems, methods, and devices described herein determine whether the subject or product is infected with a pathogen and / or generate a score representative of the subject’s or product’s health, wellness or consumability. Where a score that deviates from a set threshold value, such as a higher or lower value depending on the parameters of the threshold set, indicates that the subject or product is healthy, well, or suitable for consumption, not infected with a pathogen and / or is of low risk to others in a population or indicates that the subject or product has compromised health, is infected with a pathogen, not suitable for consumption, and / or is a risk to others in the population.

[0069]

[0068] Such a diagnostic platform may replace the complex optical systems and expensive fluorescent labels used for optical detection and the electrodes and electroactive agents used in existing electrochemical and FET techniques with common electronic components. In some aspects, the amplification can be isothermal. In some embodiments, the amplification is loop-mediated isothermal amplification (LAMP). In some embodiments, the amplification is reverse transcription loop-mediated isothermal amplification (RT- LAMP). The platform described herein is inexpensive, robust, portable, and consumes less power than traditional diagnostic systems. In some aspects, the diagnostic platform is small enough to fit in the palm of a consumer’s hand and capable of performing in the field, for example, a diagnosis in a doctor’s office, in the home, in a location remote from a medical facility.

[0070]

[0069] Many commercially available nucleic acid detection platforms utilize traditional PCR, thereby requiring temperature cycling, fluorescent labels and optical detection instrumentation. These factors result in expensive, lab-based instrumentation which employ delicate, vibration sensitive detectors, costly fluorescent markers, and have a large footprint. The equipment requires operation, and frequent calibration, by highly trained personnel.

[0071]

[0070] These large, unwieldy platforms make routine use of conventional NAAT challenging to use in the clinic, much less in the home. NAAT remains a costly and slow strategy closely tied to centralized laboratory facilities. The presently disclosed technology, in contrast, avoids these challenges.

[0072]

[0071] A hurdle to point of care (“POC”) testing is the potential inhibition of amplification by interferents often encountered in crude, unprocessed clinical samples such as cloacal matrix, whole blood, saliva, mucus, or any other bodily fluid or biological component. The mitigation of amplification inhibitors may challenge the direct detection of target nucleic acids from clinically relevant biologic samples. As described herein, a sample may comprise one or more of cloacal matrix, blood, saliva, mucus, or any other bodily fluid or biological secretion or component. Cloacal samples, in particular poultry cloacal samples such as chicken cloacal samples, can include particularly high levels of inhibitors and solid particulates that may interfere with amplification.

[0073]

[0072] Traditional detection strategies commonly rely on fluorescence detection techniques. Such techniques may be complex, more expensive, and require precision optical systems. The present disclosure, on the other hand, generally relies on electrical detection systems. Such electrical detection systems may leverage microelectronics that consume relatively low power and can be manufactured at a reduced cost due to high volume manufacturing. Thus, electrical detection of genomic material may transfer the advances of the computer industry to bioassay sensing.

[0074]

[0073] Existing electronic methods for monitoring amplification may require the binding of an electrochemically active label or the selective binding of the amplified material to a surface. However, when used in real world clinical applications, these techniques often suffer from slow response times, biofouling of the electrode or binding surfaces resulting in poor signal to noise ratios, and limitations on the lifetime and reliability of the device. While potentially enabling great sensitivity, the use of electrochemical or field effect transistor “FET” detection adds a layer of complexity to the detection. This can result in more expensive and less robust strategies than POC and other consumer applications typically dictate. Accordingly, the need for additional diagnostic devices is manifest.

[0075]

[0074] The platform disclosed herein relies on measurement of the change in electrical conductivity that occurs during nucleic acid amplification. In sum, during biochemical synthesis of DNA from nucleotide triphosphates, the number and the mobility of electrically charged molecules are altered. This, in turn, results in a change in the solution conductivity as amplification progresses. This change in solution electrical conductivity may be sensed using frequency-dependent capacitively coupled contactless conductivity detection (‘ C4D”).

[0076]

[0075] In some implementations, , / C4D uses a pair of electrodes in close proximity to, but not in contact with, a fluid disposed in an amplification chamber to measure the solution’s electrical properties. The ability to measure the properties of the solution in this way, without direct contact, avoids the challenges of surface fouling common to other electrical measurement methods.

[0077]

[0076] In some implementations, utilizing / C4D, a high frequency alternating current (“AC”) signal is applied to the excitation electrode. This signal is capacitively coupled through the solution where it is detected at the signal electrode. By comparing the excitation signal with the signal at the signal electrode, the solution’s conductivity can be determined.

[0078]

[0077] Informed by high-resolution finite element models and empirical studies, specific tolerances of , / C4D based technology may achieve the optimal detection sensitivity and dynamic sensing range for particular implementations of the platform. Such calculated and empirically determined parameters of microfluidic dimensions, capacitive coupling characteristics, and the applied frequency can enable the determination of the effective parameters for detecting solution conductivity changes. In some embodiments, the parameters corresponding to optimal detection can be interdependent variables. According to the following equation, the measured impedance is a function of the solution resistance, capacitance and the applied frequency:

[0079] Z = R - (l / pi*f*C)*j

[0080]

[0078] As the thickness of the electrode passivation layer increases, a parasitic capacitance due to this layer consequently increases. The optimal AC frequency with which to measure solution conductivity by , / C4D therefore can be chosen with respect to the capacitance of the passivation layer.

[0081]

[0079] Some embodiments provided herein include aspects disclosed in WO 2020 / 132008; WO 2016 / 057422; WO 2018 / 057647; WO 2020 / 132042; WO 2020 / 132042; WO 2020 / 132005; WO 2020 / 132010; WO 2020 / 132008; WO 2022 / 035995; U.S. 2016 / 0097740; U.S. 2016 / 0097741; U.S. 2016 / 0097739; U.S. 2016 / 0097742; U.S. 2016 / 0130639; and U.S. App. No. 18 / 612,911, which are each expressly incorporated by reference in its entirety for all purposes.

[0082] Overview of Example Cartridges. Readers, and Signal Processing

[0083]

[0080] FIGs. 1 A-1E depict an example type or format of cartridge 100 configured to detect a target, such as a nucleic acid e.g., a desired DNA or RNA sequence, which can be used in conjunction with one or more of the handheld systems disclosed herein. In some embodiments, the target may be a viral target, bacterial target, antigen target, parasite target, microRNA target, or agricultural analyte. Preferably, such targets are selected viral, bacterial, parasite, microRNA, or agricultural DNA or RNA sequences e.g., sequences complementary to selected primers designed to identify the presence or absence and / or amount of such targets. Some embodiments of the cartridge 100 can be configured for testing for the presence or absence and / or amount of a single target, while some embodiments of the cartridge 100 can be configured for testing for multiple targets, optionally simultaneously or within a short time after the first identified result. In some embodiments, the cartridge 100 may be configured to test for enzymes (for example, in the evaluating and / or analyzing for enzyme replacement therapy). In some embodiments, the cartridge 100 may be configured to test for environmental contaminants such as pesticide residues (e.g., glyphosate, and so forth), heavy metals, benzene residues, and so forth. In some embodiments, the cartridge 100 may be configured to test for or identify pathogens, genomic materials, proteins, and / or other small molecules or biomarkers. In some embodiments, the cartridge 100 may be configured to test for and / or identify elevated hormone levels, elevated cells counts, low cell counts, tumor cells, and so forth, for use in oncology applications. In some embodiments, the cartridge 100 may be configured to identify and / or test for microRNA or used to test for infections, diseases, and so forth often of concern with respect to food safety and / or plasma and / or blood screenings. In some embodiments, the cartridge 100 may be further configured to identify and / or test for rare infectious diseases, tick and / or mosquito borne (or other insect, plant, and / or animal vector borne) diseases. In some embodiments, the cartridge 100 may be configured to test for and / or identify norovirus and / or rotavirus, for example in water quality applications. In some embodiments, the cartridge 100 is configured to test for anything any of the other cartridges described herein test for, and vice versa. The cartridge 100 includes, among other components, a cartridge body 102, a cover 114, electrodes 104, and a cap 110. These components will be described in further detail below.

[0084]

[0081] Referring now to FIGs. 1 A-l E, the cartridge body 102 may be thermoformed from a polyethylene or similar plastic material. The cartridge body 102 may house various components and / or features of the cartridge 100. For example, the cartridge body 102 may house at least a portion of the electrodes 104, a sample receptacle 142, a sample mixing and microfiltration region 130 (described further below), a reagent stored in a reagent blister 140, and an integrated reagent blister rupture feature 144, a distribution tree 128 for a sample / reagent mixture from the sample mixing and microfiltration region 130, an exhaust port 126 to vent gases, a degassing gas-permeable membrane 120, and a plurality of reaction wells 122 configured to allow the electrodes 104 to generate signals based on the sample / reagent mixture. The cartridge body 102 also includes a thumb detent configured to be grasped by a user to facilitate removal of the cartridge 100 from the analyzer, described in further detail below. The cartridge body 102 also, optionally includes an isolation cap 110 or a locking isolation cap 110 configured to isolate, close, or protect the sample receptacle 142 from external variables. The cover 114 of the cartridge body 102 may be thermoformed from a plastic or similar material and includes an exhaust port crush valve 106 and well isolations crush valves 112.

[0085]

[0082] The well isolations crush valves 112 may isolate fluid within the plurality of reaction wells 122. In some embodiments, the reaction wells 122 may function similarly to the test wells 1258 described above. The electrodes 104 may electrically connect electrodes at the reaction wells 122 to circuitry of a reader device described further below.

[0086]

[0083] The sample receptacle 142 can receive a sample. In some examples, the sample receptacle 142 can receive a sample on a swab head. In some embodiments, the sample receptacle 142 includes a tapering, tubular portion 148, a scraper portion 150, dispensing portion 152, and the isolation cap 110. In some examples, the isolation cap 110 can lock to the sample receptacle 142. The isolation cap 110 and the sample receptacle 142 are configured to fit together to form a sealed or substantially sealed swab assembly. The tapering, tubular portion 148 includes a tapering tube channel sized and shaped to receive a swab including mucus or another sample and to be sealed or substantially sealed by the isolation cap 110. The reagent blister 140 may contain one or more reagents (for example, a liquid reagent, a buffer, etc.) during shipping and handling before the cartridge 100 is inserted into the corresponding analyzer. In some embodiments, the reagent blister 140 includes only reagents in a liquid form. In some embodiments, the reagent blister may comprise a sealed compartment, for example sealed with a foil seal or the like, similar to the sealed channel 1226 described above. The sample mixing and macrofiltration region 130 may be configured and operate to facilitate mixing of fluids and so forth therein (for example, the sample with the reagent). In some embodiments, the tapering, tubular portion 148 may further include the scraper 150 configured to facilitate acquisition of the sample from the swab inserted into the sample receptacle 142. Further details are provided below. The cartridge body 102 includes snap-fit clips 146 that may lock and / or engage with snap-fit openings to interlock and hold the cartridge body 102 together.

[0087]

[0084] The cartridge 100 may be a disposable cartridge that is fully integrated, enables detection of one or more pathogens, and includes no moving parts, improving reliability of the cartridge 100.

[0088]

[0085] The cartridge 100 may be used in conjunction with an analyzer, similar to the analyzer or reader device (for example, reader device 300). In some embodiments, the analyzer (also referred to herein as the “reader device”) may be handheld and battery operated and enable wireless communication with an application operating, for example, on a user’s mobile phone or other mobile device. In some embodiments, the application may provide the operator with a view of the results from the analysis of the cartridge. In some embodiments, the application may communicate with a cloud storage system (or similar storage) to store data from the application that is received from or via the analyzer. In some embodiments, the analyzer, application, and cloud storage may enable secure communications and may aggregate information from various cartridge samples to enable treatment decisions (for example, identify that a sample being tested with a cartridge indicates a particular sickness, etc., based on a comparison of the analysis results from the analyzer with results from a historical database of analyses and corresponding sickness, etc., determinations. For example, if the cartridge analyzed by the analyzer indicates influenza, as compared to similar results in the cloud-based historical database, the application may identify appropriate treatments or therapies for influenza and present them to the operator of the application (e.g., to the veterinarian, the physician, or health care practitioner, etc.). In some embodiments, the data stored in the cloud may be analyzed in real time to identify outbreaks of diseases, and so forth. Additionally, such information may be used to update manufacturers of vaccines and medications in response to the outbreaks, etc., to ensure sufficient stockpiles of vaccines and / or medications are available.

[0089]

[0086] In some aspects, a cartridge in accordance with the present disclosure can include a filter for separating solid particles from a fluid sample. Such cartridges may additionally include any of the features discussed with reference to FIGs. 1 A-1K. FIGs. 2A and 2B diagram an example cartridge 200 including a plug filter 204. FIG. 2A presents a transparent view of the cartridge 200. FIG. 2B presents a cross-sectional view of cartridge 200. The plug filter 204 may be positioned between the sample receptacle 202, and a fluidic path 206. Sample fluid introduced to the sample receptacle 202 can flow through the plug filter 204 prior to flowing through the fluidic path 206 toward a mixing chamber and / or an analysis chamber of the cartridge 200. The cartridge may include a cap 208 for sealing the sample receptacle 202.

[0090]

[0087] FIGs. 2C and 2D illustrate another example cartridge 250 including a filter 208. FIG. 2C provides a side view of the cartridge 250. The cartridge 250 includes a filter 208 that covers the sample receptacle 202. The cartridge includes a pipette adaptor 212 for coupling to a pipette 214. As shown in FIG. 2D, the filter 208 is positioned between a cavity of the pipette adaptor 212 and the sample receptacle 202, such that the fluid introduced by the pipette 214 to the cartridge 250 flows through the filter 208 before reaching the sample receptacle 202. From the sample receptacle 202, the filtered fluid 210 can flow to mixing chambers via one or more of the fluidics channels 206.

[0091]

[0088] The filters 204 or 208 can include pores of suitable size to filter out solid particles from a fluid sample while allowing passage of target molecules (e.g., nucleic acids) from the fluid sample. The filter 1306 can include pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The filters 204 or 208 can include pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the pore sizes are from 20 to 50 pm in diameter.

[0092]

[0089] FIGs. 3A and 3B depict an example of the other handheld analyzer system 300 disclosed herein. The analyzer 300 includes the features described above of other analyzers, including a slot for insertion of the cartridge, for example the cartridge 100. In some embodiments, the analyzer 300 comprises a printed circuit board (PCB) 302 on which various electrical components are disposed and electrically connected. The PCB 302 includes a communication or charging port, such as a USB (or similar) port 304. The PCB 302 further includes a microprocessor 306 and a digital signal processor (DSP) 308. The PCB 302 further comprises an analog sub-section 310 and cartridge mating connectors 312 to electronically couple to the cartridge 100. The PCB 302 further comprises a cryptographic processor 314, a battery controller 316, and storage 318. The PCB 302 may also include one or more mounting holes or devices 320.

[0093]

[0090] Operation of the cartridge 100 with the analyzer 300 is now described. Before operating the cartridge and the analyzer 300, the application may be used to select a particular cartridge 100 that will be used with the analyzer 300. In some embodiments, the selection may comprise indicating one or more parameters associated with the particular cartridge 100 to the analyzer 300. A swab may be used to collect a fluid sample (for example, a cloacal matrix, nasal fluid, or sputum sample). The swab with the fluid sample from the subject may be pressed into the sample receptacle 142 until the swab stops moving into the sample receptacle 142. As described above, the sample receptacle 142 tapers down in diameter to a shape / size that compresses the bristles or other material of the swab against the walls of the sample receptacle 142. In some embodiments, the sample receptacle 142 comprises one or more walls (or other surfaces) molded with a specifically selected surface finish. The surface finish enables wetting the fluid sample to the walls. As the swab continues to its fully compressed position as it is pushed into the sample receptacle 142, the fluid that has been wetted to the walls is forced further in front of the swab because of the decreasing crosssection profile of the sample receptacle 142. As the swab is removed, the fluid that is forced ahead of the swab is drawn back with the swab being removed, trailing behind the swab. At the point in which the bristles or material of the swab have separated far enough from the walls of the sample receptacle 142, a surface tension of the fluid sample that is wetted to the walls is sufficient to retain at least a portion of the fluid sample in the sample receptacle 142 (for example, a reservoir portion of the sample receptacle 142). Thus, the fluid sample to be analyzed by the analyzer is removed from the fluid sample initially collected on the swab and remains in place as the cap 110 is closed, sealing or substantially sealing the sample receptacle 142 and the cartridge 100.

[0094]

[0091] In some embodiments, a user may insert a swab 160 into the sample receptacle 142 of the cartridge 100 as shown in, for example, FIG. 1G. Once inserted into the sample receptacle 142, the swab 160 may be broken or cut, for example, at 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10mm or within a range defined by any two of the aforementioned sizes below a swab stopper 162 or the cartridge interface, such that two separate pieces are generated one being a portion of the swab 160 including a sample (e.g. a cloacal sample), which remains inside the sample receptacle 142 and a portion of the swab 160 without the mucus or another type of sample (for example, a second portion), which is separated from the first portion and discarded. FIG. 1H schematically illustrates a location 170 where swab 160 can be broken or cut (for example, snapped) into two separate pieces. As shown in FIG. II, the cap 110 can be moved to close (for example, seal) the sample receptacle 142 of the cartridge 100, leaving the first portion of the swab (for example, a portion with mucus or another type of sample) positioned inside the sample receptacle 142. After the sample receptacle 142 of the cartridge 100 is closed via the cap 110, a user (for example, a care provider, an operator of the analyzer 300 and the cartridge 100) can insert the sealed cartridge 100 into the analyzer 300 as shown in FIG. 1 J to test the collected sample.

[0095]

[0092] In some embodiments, as shown in FIG. IF, a swab 160 can include a stopper 162. The stopper 162 can be a radial protrusion from a shaft of the swab 160 that can cover an opening of the sample receptacle 142 when the swab 160 is inserted into the sample receptacle 142. As such, when the swab 160 is inserted into the sample receptacle 142, the stopper 162 can prevent the swab 160 from being inserted into the sample receptacle 142 further, for example, than a predetermined distance.

[0096]

[0093] The stopper 162 can indicate where to break or cut the swab 160 after inserting the swab 160 into the sample receptacle 142. In some embodiments, the swab 160 has a perforated or marked section 164, which indicates a position to break or cut the swab 160. When the stopper 162 covers the opening of the sample receptacle 142, a user can break or cut a shaft of the swab 160, for example, proximal to or against the opening of the sample receptacle 142 using the stopper 162 as a lever or guide for the cut In some examples, the stopper 162 can remain with, for example, a portion of the swab 160 with the bristles or flock. In other examples, the stopper 162 may break off from the portion of the swab 160 with the bristles or flock and be discarded.

[0097]

[0094] In some embodiments, the location of the stopper 162 can be such that the bristles or flock (or a portion of the swab 160 having mucus or another type of sample) of the swab 160 are positioned near or adjacent to the scraper 150 when the stopper 162 stops the swab 160 from entering further into the sample receptacle 142. Alternatively, the location of the stopper 162 can be such that the swab 160 is in its fully compressed or substantially fully compressed position in the sample receptacle 142 when the stopper 162 stops the swab 160 from moving further into the sample receptacle 142. Alternatively, the location of the stopper 162 can be such that, for example, the bristles or flock (or a portion of the swab 160 having mucus or another type of sample) of the swab are positioned further down into the sample receptacle 142 past the scraper 150. The stopper 162 can desirably indicate to users how far the swab 160 is preferably inserted into the sample receptacle 142 to allow the cartridge 100 via the scraper 150 to collect mucus or other types of sample from the swab 160.

[0098]

[0095] In some embodiments, the sample receptacle 142 can include, for example, a retainer 180 that can hold a swab once it is inserted into the sample receptacle 142. For example, the retainer 180 can include fingers 182 that can function as a clamp for the swab 160. After the swab 160 is inserted into the sample receptacle 142, the swab 160, for example, pressed against the retainer 180 such that the swab 160 can be positioned between the fingers 182 and held in place in the sample receptacle 142. Once the swab is held in place in the sample receptacle 142, a user (for example, a care provider, an operator of the analyzer 300 and the cartridge 100) can insert the cartridge 100 into the analyzer 300 to test the sample from the swab. Other designs of the retainer 180 suitable to hold, for example, the shaft of the swab 160 may be used in conjunction with the sample receptacle 142. Having the retainer 180 to hold the swab 160 in place inside the sample receptacle 142 can desirably eliminate the need to break the swab and closing the cap 110 to keep the swab in place inside the sample receptacle 142 during testing.

[0099]

[0096] In other examples, a fluid sample may be added to the sample receptacle 142 without a swab. As an illustrative example, a fluid sample can be pipetted into the sample receptacle 142. In some examples, buffer may be added to such a fluid sample prior to the sample being introduced to the sample receptacle 142. In some examples, buffer may be added to the fluid sample while it is in the sample receptacle 142.

[0100]

[0097] The reagent blister 140 and the integrated rupture feature 144 of the cartridge 100 can be used to improve testing of collected samples. Prior to receiving samples (for example, mucus or other types of samples), the integrated rupture feature 144 may be pressed. When the integrated rupture feature 144 is pressed, it may rupture and release solution or solutions stored inside (that is, inside the integrated rupture feature 144) e.g., 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 microliters of solution or an amount of solution that is within a range defined by any two of the aforementioned volumes, which then enters the sample receptacle 142. Once sample is collected via the sample receptacle 142, the cap 110 can be closed to seal the sample receptacle 142. Once the cap 110 is closed, the reagent blister 140 may be pressed. The pressing of the reagent blister 140 can desirably mix the solution (for example, a buffer) released from the rupture feature 144 with solution (for example, reagent) previously stored in the reagent blister 140. Additionally, the pressing of the reagent blister 140 can cause the mixed solution to flow towards, for example, the sample mixing and macrofiltration region 130 of the cartridge 100. The mixing of the solutions from the integrated rupture feature 144 and the reagent blister 140 can provide improved recovery of sample from the swab 160, improved reaction rate between the collected sample and the reagent, and improved test results (for example, decreased rate of false-positive). In some embodiments, after the solution from the integrated rupture feature 144 is provided to the sample receptacle 142, the swab 160 is inserted into the sample receptacle 142 and is twisted or rotated a plurality of times e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times or an amount that is within a range defined by any two of the aforementioned times.

[0101]

[0098] After the sample receptacle 142 of the cartridge 100 is closed via the cap 110, the operator of the analyzer and cartridge may be prompted to insert the now sealed cartridge 100 into the analyzer 300. The process of inserting the cartridge 100 into the analyzer 300 may actuate multiple features on the cartridge 100 using static and / or passive components. For example, the singular action of inserting the cartridge 100 into the analyzer 300, as performed by the operator, can have its functionality divided into 3 major phases, with some overlap amongst them, as described below.

[0102]

[0099] The first major phase involves inserting the cartridge 100 into the analyzer 300. As the cartridge 100 is first inserted into the analyzer 300, a static platform located on the bottom mating surface of the cartridge / analyzer interface in the analyzer 300 depresses a living hinge via interference (for example, mechanical interference). Thus, the static platform translates a linear motion of the cartridge 100 as it is inserted into the analyzer 300 into an angular displacement of a lever mechanically coupled to and engaged with the living hinge. The displacement of the lever / living hinge may result in the rupture of the reagent blister 140. In some embodiments, the lever and / or living hinge may correspond to the integrated rupture feature 144, introduced above. The lever coupled to the living hinge may continue to rotate about the living hinge and flex along a length of the lever to depress the reagent blister 140, thereby distributing the reagents into the cartridge 100 (for example, into the sample mixing and macrofiltration region 130 of the cartridge 100). In some embodiments, forces associated with this distribution action may cause the sample in from the sample receptacle 142 to mix with and be diluted by the reagents from the ruptured reagent blister 140 in the sample mixing and macrofiltration region 130. The forces may further distribute the mixed sample and reagent to the reaction wells 122, rehydrating dried and / or lyophilized reagents disposed in the reaction wells 122. The flow of the mixed sample and reagent may terminate at the exhaust gas-permeable membrane 124 located at the exit of each reaction well 122, thereby ensuring uniform filling in each well. All of these processes may occur before the cartridge 100 is inserted into the analyzer 300.

[0103]

[0100] As a second of the three major phases, the reaction wells 122 are isolated. As the mixed sample and reagent is distributed through the cartridge 100 as described above, a path through with the mixed sample and reagent flows transitions from the distribution tree 128 of the injection molded cartridge body 102 (for example, the portions of the cartridge 100 not including and below the thermoformed cover 114) into small channels formed in the thermoformed cover 114. In some embodiments, the thermoformed cover 114 is attached to the cartridge body 102 via pressure-sensitive adhesive (PSA) above the plane of the injection molded cartridge body 102. In some embodiments, the thermoformed cover 114 is translucent. In some embodiments, the thermoformed cover 114 also serves as a housing for or includes other fluidic channel components for the cartridge body 102. In some embodiments, these other fluidic channel components, for example, the well isolation crush valves 112, act as isolation valves for each individual reaction well 122. As the cartridge 100 nears the end of its insertion into the analyzer 300, after each of the reaction wells 122 has been filled with the mixed sample and reagent, passive features located on the upper mating surface of the analyzer 300 “crush” the isolation crush valves 112 (which may comprise thin channels) down against the PSA bonding the thermoformed cover 114 or a film to the injection molded cartridge body 102. Because of the nature of the PSA, the PSA conforms to the deformed shape of the thin film channel in the cartridge body 102, serving to isolate the reaction well 122 fluidically from the channels that join the reaction wells 122 together fluidically. This isolation may prevent crosstalk and diffusion from reaction well 122 to reaction well 122. Additionally, this isolation may serve to isolate the reagents and the byproducts of the reaction from the analyzer 300 and user.

[0104]

[0101] While the cartridge 102 is being inserted into the analyzer 300, the third phase occurs. As described above, the well-isolation crush valves 112 may protrude from an upper surface of the thermoformed cover 114 on the cartridge body 102 so that they are crushed when the cartridge 102 is inserted into the analyzer 300. This orientation combined with a dual sided heating design for the cartridge 100 in the analyzer 300 necessitated a method of removing a heater surface out of the way of the exhaust crush valve 106. In order to passively accomplish this, an upper heater is mounted to an integrated crossbar and leaf spring of the analyzer 300. In some embodiments, two cam runners are positioned in a cartridge receptacle of the analyzer 300 such that when the cartridge 100 begins to be inserted, the cam runners push the upper heater up and out of the way of the crush valves (for example, the well isolation crush valves 112 and / or the exhaust port crush valve 106) disposed on the thermoformed cover 114. The cam runners continue to pass along an upper edge of the cartridge body 102 until the cam runners arrive at a molded drop in the upper edge of the cartridge body 102. The molded drop in the upper edge of the cartridge body 102 may be timed or positioned in such a way so as to allow the upper heater to press back down against the cartridge body 102 while not interfering with the thermoformed valves on the thermoformed cover 114. In some embodiments, the cam runners dropping into the molded drop of the cartridge body 102 may also double as a retention feature for the cartridge 100 in the analyzer 300. As such, the user may displace a leaf spring that moves the upper heater up and out of the way of the crush valves and displacing the cam runners from the molded drop in order to extract the cartridge 100 after the testing is completed in the analyzer 300.

[0105]

[0102] After these three major phases are completed, the cartridge 100 is fully inserted into the analyzer 300, the mixed sample and reagent are distributed to the reaction wells 122 and isolated in each reaction well 122. The upper heater in the analyzer 300 may uniformly contact the upper surface of the cartridge body 102, sandwiching the cartridge body 102 against a lower heater in the analyzer 300. Upon full insertion, the cartridge 100 may also establish an electrical connection to the analyzer 300 via the flexible electrode layer 104, thereby enabling the analyzer 300 to begin a test of the mixed sample and reagent in the reaction wells 122.

[0106]

[0103] In some embodiments, the method for using the cartridge 100 with the analyzer 300 to detect a target involves first collecting a sample fluid from a user. In some embodiments, the sample fluid may be collected using a swab or other similar sample collecting method or device. Once the sample fluid is collected, the sample fluid is introduced into the cartridge 100. For example, when the sample fluid is collected using the swab, the swab is inserted into the cartridge 100 via the sample receptacle 142. The optional cartridge cap 110 may be closed and the cartridge 100 may be inserted into the analyzer 300. The insertion of the cartridge 100 may cause a reagent blister 140 to rupture and mix the reagents contained therein with the sample fluid. In some embodiments, the mixture of the reagents and the sample fluid is conveyed to the reaction wells 122. The reaction wells 122 may each have a volume of approximately 25 microliters (pL). In some embodiments, the reaction wells 122 has a volume, size, and / or shape that is based on the overall volume of liquid which will fill the reaction wells 122 as well as a favorable geometry above the sensing electrode. For example, in some embodiments the reaction wells 122 are circular, triangular, or rectangular, or any other polygonal shape. In some embodiments, when the reaction wells 122 comprise a plurality of reaction wells (for example, eight (8) reaction wells), the electrodes 104 (or corresponding circuitry and / or circuit board components / parameters) of the cartridge 100 may enable the analyzer 300 to simultaneously test each of the reaction wells 122 (for example, each representing a different channel) at a plurality of frequencies (for example, three (3) frequencies). In some embodiments, each reaction well 122 has a depth or height of approximately 1 millimeter (mm). In some embodiments, the reaction wells 122 may have a shape such that the volume of each reaction well 122 is or is approximately 25 pL when each reaction well 122 has a depth or height of 1 mm. In some embodiments, the reaction wells 122 contain dried and / or lyophilized enzymes and primers. The primers and enzymes may be spotted into the reaction wells 122 as liquids and dried and / or lyophilized (for example, in two spots per well). Thus, the reaction wells 122 may include dried and / or lyophilized primers and enzymes while the reagent blister 140 may include the wet or liquid reagents (e.g., buffer fluid). In some embodiments, the reagent blister 140 and / or the reaction wells 122 may contain liquid reagents and / or primers and enzymes, respectively, configured to allow for the testing of one or more of a viral target, a bacterial target, an antigen target, a parasite target, a microRNA target, an agricultural analyte, an enzyme, an environmental contaminant, pathogens, genomic materials, proteins, PSA levels, elevated or reduced cell counts, specific cells and / or cell types, infections and / or diseases associated with a particular industry or environment, infectious diseases, vector borne diseases, norovirus, rotavirus, and / or any other small molecules or biomarkers. In some embodiments, the dried and / or lyophilized enzymes and primers are contained in the reagent blister 140 or elsewhere in the cartridge 100. Once the mixture of the reagents and the sample fluid is introduced to the reaction wells 122, one or more heating elements in the analyzer 300 are activated to increase a temperature of the mixture in the reaction wells 122 and impedance sensors begin tracking data in real time. The analyzer 300 then conducts a test through an isothermal nucleic acid amplification process, and any results are communicated to an application and / or a database for further analysis.

[0104] In some embodiments, a reader device, such as the reader device 300 may incorporate additional functions and / or components in accordance with the present disclosure.

[0107]

[0105] Similarly, the cartridge 100 may incorporate additional functions and / or components discussed herein. Similarly, the cartridge 100 may be processed by any reader device described herein.

[0108]

[0106] In some aspects, a system for detecting a target in a sample includes a removable fluidics cartridge that is couplable to a companion reader device. A user can apply a sample to the cartridge and then insert it into the reader device. The reader device is configured for performing the testing procedures using the cartridge and analyzing the test data to determine the presence, absence, or quantity of a target in the sample. For example, the cartridge can be provided with the desired agents, proteins, or other chemical matter for an amplification process by which a target initially present in the sample is amplified. Specifically, some cartridges can be provided with the desired chemical matter for nucleic acid testing, wherein genomic material in the sample is exponentially copied using a molecular amplification process, as described herein. The cartridge can also include a test well for containing the amplification process, where a test well refers to a well, chamber, channel, or other geometry configured for containing (or substantially containing) test fluid and constituents of the amplification process. The reader device may maintain a desired temperature or other test environment parameters for the cartridge to facilitate the amplification process and can electronically monitor a test well of the cartridge throughout some or all of the amplification process. The reader device can thus gather signal data representing the impedance of the test well over time during the amplification process and can analyze the impedance as described herein to ascertain the presence, absence, or quantity of the target in the sample. As an example, the amplification process can range from five minutes to sixty minutes, with some examples ranging from ten minutes to thirty minutes. Preferably, in some embodiments, the amplification products are detected while being suspended in the fluid within the wells such that the amplification products are not attached or sequestered to the wells or fixed or bound to probes, which are bound to the wells. In other embodiments, the amplification products are detected as they are attached or sequestered to the wells e.g., fixed or bound to probes, which are bound to the wells.

[0107] Such systems can beneficially provide target detection performable in a clinical setting or even the home of a user, rather than requiring the sample to be sent to a laboratory for amplification and analysis. In the clinical setting, this can avoid the delays of conventional nucleic acid testing thereby enabling clinicians to determine diagnoses within relatively short times. As such, the disclosed systems enable clinicians to rapidly develop treatment plans, rather than requiring the clinician to wait for hours or even days to receive test results back from a laboratory. Particularly when used to diagnose pathologies that progress quickly, the disclosed systems can avoid the delays associated with laboratory testing that can negatively impact the treatment and outcome of the subject.

[0109]

[0108] As another benefit, the disclosed systems can be used outside of the clinical setting (e.g., in the field, in rural settings without easy access to an established healthcare clinic) to detect health conditions such as contagious diseases (e.g., Ebola), thus enabling the appropriate personnel to take immediate action to prevent or mitigate the spread of a contagious disease. Similarly, the disclosed systems can be used in the field or at the site of a suspected hazardous contaminant (e.g., anthrax) to quickly determine whether a sample contains the hazardous contaminant, thus enabling the appropriate personnel to take immediate action to prevent or mitigate human exposure to the contaminant. Additionally, the disclosed systems can be used to detect contaminants in the blood or plasma supply or in the food industry. It will be appreciated that the disclosed systems can provide similar benefits in other scenarios in which real-time detection of a target enables more effective action than delayed detection through sending a sample to an off-site laboratory.

[0110]

[0109] Another benefit of such systems is their use of low-cost, disposable single use cartridges together with a reusable reader device that can be used many times with different cartridges and / or for tests with different targets. In some embodiments disclosed herein, a single use cartridge includes a cartridge body and a cap which, when mechanically coupled together, create pressurized air that propels a collected sample from the cap into a mixing well and a test well of the cartridge body, reducing a necessary level of skill required to operate the reader device and reducing the complexity of both the cartridge and the reader device.

[0111]

[0110] FIGs. 4A-4N depict various examples of electrode configurations that can be used in a test well of the cartridge of FIGs. 1A-1D or in the test well or channel of another suitable target detection cartridge as described herein. The test wells shown in FIGs. 4A-4N are depicted as circular, however the electrodes can be used in test wells of other geometries in other examples. Unless otherwise noted, the solid circles in FIGs. 4A-4N represent contacts between the disclosed electrodes and conductors leading to or from the electrode. “Width” as used below refers to a dimension along the horizontal direction of the pages of FIGs. 4A-4N, and “height” as used below refers to a dimension along the vertical direction of the pages of FIGs. 4A-4N. Though depicted in a particular orientation, the illustrated electrodes of FIGs. 4A-4N can be rotated in other implementations. Further, the disclosed example dimensions represent certain potential implementations of the electrode configurations 400A-400G, and variations can have different dimensions that follow the same ratios between the provided example dimensions. The electrodes shown in FIGs. 4A- 4N can be made from suitable materials including platinum, gold, steel, or tin. In experimental testing, tin and platinum performed similarly and suitably for certain test setups and test targets.

[0112]

[0111] FIG. 4A depicts a first electrode configuration 400A wherein the first and second electrodes 405 A, 405B are each formed as a semicircular perimeter. The straight edge of the first electrode 405A is positioned adjacent to the straight edge of the second electrode 405B and separated by a gap along the width of the configuration 400 A. The gap is larger than the radius of the semicircle of the electrodes. Thus, the first and second electrodes 405A, 405B are positioned as mirrored semicircular perimeters. In one example of the first electrode configuration 400 A, the gap between the closest portions of the first and second electrodes 405A, 405B spans approximately 26.369 mm, the height (along the straight edge) of each of the electrodes 405 A, 405B is approximately 25.399 mm, and the radius of the semicircle of each of the electrodes 405A, 405B is approximately 12.703 mm.

[0113]

[0112] FIG. 4B depicts a second electrode configuration 400B. Similar to the first electrode configuration 400A, the first and second electrodes 410A, 410B of the second electrode configuration 400B are each formed as a semicircular perimeter and are positioned as mirrored semicircles with their straight edges facing one another. The first and second electrodes 410A, 410B of the second electrode configuration 400B can be the same size as the first and second electrodes 405 A, 405B of the first configuration 400A. In the second electrode configuration 400B, the gap along the width of the configuration 400B between the first and second electrodes 41 OA, 41 OB is smaller than in the first configuration 400A, and the gap is smaller than the radius of the semicircle of the electrodes 410A, 41 OB. In one example of the second electrode configuration 400B, the gap between the closest portions of the first and second electrodes 41 OA, 41 OB spans approximately 10.158 mm, the height (along the straight edge) of each of the electrodes 410A, 41 OB is approximately 25.399 mm, and the radius of the semicircle of each of the electrodes 41 OA, 41 OB is approximately 12.703 mm.

[0114]

[0113] FIG. 4C depicts a third electrode configuration 400C having first and second linear electrodes 415A, 415B separated by a gap along the width of the configuration 400C, where the gap is approximately equal to the height of the electrodes 415 A, 415B. The width of the electrodes 415A, 415B is approximately one half to one third of the height of the electrodes. In one example of the third electrode configuration 400C, the gap between the closest portions of the first and second electrodes 415 A, 415B spans approximately 25.399 mm, the height of each of the electrodes 415 A, 415B is also approximately 25.399 mm, and the width of each of the electrodes 415 A, 415B is approximately 10.158 mm. The ends of the first and second electrodes 415A, 415B can be radiused, for example having a radius of around 5.078 mm.

[0115]

[0114] FIG. 4D depicts a fourth electrode configuration 400D having first and second rectangular electrodes 420A, 420B separated by a gap along the width of the configuration 400D, where the gap is approximately equal to the width of the electrodes 420A, 420B. In one example of the fourth electrode configuration 400D, the gap between the closest portions of the first and second electrodes 420A, 420B spans approximately 20.325 mm, the height of each of the electrodes 420A, 420B is also approximately 23.496 mm, and the width of each of the electrodes 420 A, 420B is approximately 17.777 mm.

[0116]

[0115] FIG. 4E depicts a fifth electrode configuration 400E having first and second linear electrodes 425A, 425B separated by a gap along the width of the configuration 400E, where the gap is approximately equal to the height of the electrodes 425A, 425B. The fifth electrode configuration 400E is similar to the third electrode configuration 400C, with the width of the electrodes 425 A, 425B reduced to around one half to two thirds of the width of the electrodes 415A, 415B while having the same height. In one example of the fifth electrode configuration 400E, the gap between the closest portions of the first and second electrodes 425 A, 425B spans approximately 25.399 mm, the height of each of the electrodes 425A, 425B is also approximately 25.399 mm, and the width of each of the electrodes 425 A, 425B is approximately 5.078 mm. The ends of the first and second electrodes 425A, 425B can be radiused, for example having a radius of around 2.542 mm.

[0117]

[0116] FIG. 4F depicts a sixth electrode configuration 400F having concentric annular electrodes 430A, 430B. The inner electrode 430B can be a disc or circular-shaped electrode and can be positioned in the center of the test well. The outer electrode 430A can be a semicircular electrode formed concentrically around the inner electrode 43 OB and separated from the inner electrode 43 OB by a gap. In the sixth electrode configuration 400F, the gap is approximately equal to the radius of the inner electrode 430B. A break in the semicircle of the outer electrode 430 A occurs where a conductive lead connects the inner electrode 43 OB to the current providing conductor. In one example of the sixth electrode configuration 400F, the gap between the inner edge of the annular first electrode 430A and the outer perimeter of the circular second electrode 43 OB spans approximately 11.430 mm, the radius of the circular second electrode 43 OB is approximately 17.777 mm, and the thickness of the annulus of the annular first electrode 430A is approximately 5.080 mm. The ends of the first electrode 430A can be radiused, for example having a radius of around 2.555 mm, and the gap between the open ends of the annulus of the first electrode 435 A can be around 28.886 mm from vertex to vertex.

[0118]

[0117] FIG. 4G depicts a seventh electrode configuration 400G having concentric annular electrodes 435 A, 435B. Similar to the embodiment of FIG. 4F, the inner electrode 435B can be a disc or circular-shaped electrode having the same radius as inner electrode 430B and can be positioned in the center of the test well. The outer electrode 435 A can be a semicircular electrode formed concentrically around the inner electrode 435 A and separated from the inner electrode 435 A by a gap. In the seventh electrode configuration 400G, the gap is greater than the radius of the inner electrode 435B, for example two to three times greater. Correspondingly, the outer electrode 435B has a larger radius than the outer electrode 430B. In one example of the seventh electrode configuration 400G, the gap between the inner edge of the annular first electrode 435 A and the outer perimeter of the circular second electrode 435B spans approximately 24.131 mm, the radius of the circular second electrode 435B is approximately 17.777 mm, and the thickness of the annulus of the annular first electrode 435A is approximately 5.080 mm. The ends of the first electrode 435 A can be radiused, for example having a radius of around 2.555 mm, and the gap between the open ends of the annulus of the first electrode 435 A can be around 46.846 mm from vertex to vertex.

[0119]

[0118] In the embodiments of FIGs. 4A-4E, either electrode can be used as the excitation electrode and the other electrode can be used as the signal electrode. In the embodiments of FIGs. 4F and 4G, the inner electrode 430B, 435B is configured to be used as the excitation electrode (e.g., coupled to a current source) and the outer electrode 430A, 435 A is configured to be used as the signal electrode (e.g., provides its signal to a memory or processor). In some example tests, the sixth electrode configuration 400F exhibited the best performance of the configurations shown in FIGs. 4A-4G.

[0120]

[0119] FIGs. 4H-4N depict further examples of electrode configurations suitable for implementing three- terminal sensing and / or four-terminal sensing. In some embodiments, three-terminal sensing (e.g., potentiostat-type or 3 -wire measurement) or four-terminal sensing (e.g., Kelvin-type or 4- wire measurement) may improve the accuracy of impedance measurements in the systems and methods described herein. For example, the excitation electrode and the signal electrode may themselves carry some charge. Additionally, there may be some additional impedance related to surface effects at the electrode-fluid interface. Accordingly, a third electrode or a third and fourth electrode (e.g., a second electrode pair) may further be disposed within the test well. The third and / or fourth electrodes can carry a substantially smaller or negligible current relative to the current carried by the excitation and signal electrodes. The third and / or fourth electrodes may thus be used to accurately determine a voltage (e.g., a voltage between the third and fourth electrodes, or a voltage between the third electrode and the excitation or signal electrode). This precisely measured voltage may be used to determine an impedance measurement having enhanced accuracy. It will be understood that the configurations of three or four electrodes illustrated in FIGs. 4H- 4N are merely examples of a number of three- or four-terminal configurations that may be provided within the test wells of the present disclosure.

[0121]

[0120] FIG. 4H depicts an electrode configuration similar to the electrode configuration of FIG. 4A, with the addition of a third electrode 440A and a fourth electrode 440B disposed between the first electrode 405A and the second electrode 405B. Either or both of the third electrode 440A and the fourth electrode 440B may be used to implement three- or four-terminal sensing.

[0122]

[0121] FIG. 41 depicts an electrode configuration similar to the electrode configuration of FIG. 4B, with the addition of a third electrode 440A disposed between the first electrode 41 OA and the second electrode 41 OB. The third electrode 440A may be used to implement three-terminal sensing.

[0123]

[0122] FIG. 4 J depicts an electrode configuration similar to the electrode configuration of FIG. 4C, with the addition of a third electrode 440A and a fourth electrode 440B disposed between the first electrode 415A and the second electrode 415B. Either or both of the third electrode 440A and the fourth electrode 440B may be used to implement three- or four-terminal sensing.

[0124]

[0123] FIG. 4K depicts an electrode configuration similar to the electrode configuration of FIG. 4D, with the addition of a third electrode 440A disposed between the first electrode 420A and the second electrode 420B. The third electrode 440A may be used to implement three-terminal sensing.

[0125]

[0124] FIG. 4L depicts an electrode configuration similar to the electrode configuration of FIG. 4E, with the addition of a third electrode 440A disposed between the first electrode 425A and the second electrode 425B. The third electrode 440A may be used to implement three-terminal sensing.

[0126]

[0125] FIG. 4M depicts an electrode configuration similar to the electrode configuration of FIG. 4F, with the addition of a third electrode 440A disposed between the outer electrode 43 OA and the inner electrode 43 OB. The third electrode 440A may be used to implement three-terminal sensing.

[0127]

[0126] FIG. 4N depicts an electrode configuration similar to the electrode configuration of FIG. 4G, with the addition of a third electrode 440A and a fourth electrode 440B disposed between the outer electrode 435 A and the inner electrode 435B. Either or both of the third electrode 440A and the fourth electrode 440B may be used to implement three- or four-terminal sensing.

[0128]

[0127] FIG. 5 A schematically depicts a first electrode or excitation electrode and a second electrode or signal electrode that may be spaced apart from one another within a test well or channel of a suitable target detection cartridge as described herein.

[0128] The formation of an aggregate, nucleic acid complex, or polymer, for example during an amplification process in the test wells of cartridges in accordance with the present disclosure, can affect waveform characteristics of one or more electrical signals that are sent through a channel. As shown in FIG. 5 A, a first electrode or excitation electrode 510A is spaced apart from a second electrode or sensing electrode 51 OB within test well 505. The test well 505 can contain a test solution undergoing an amplification process. During some of all of that process, an excitation voltage 515 can be provided to the excitation electrode 510A, from which the excitation voltage 515 is transmitted into the fluid (preferably all or substantially all liquid) within the well 505.

[0129]

[0129] After passage through and attenuation by the liquid sample (represented schematically by the resistance R and reactance X), the attenuated excitation voltage is sensed or detected at the sensing electrode 51 OB. The fluid acts as a resistor R in series with the excitation electrode 510A and the sensing electrode 51 OB. The fluid also acts as in series capacitor(s), shown by the reactance X. The raw sensed signal during some or all of the duration of a test can be represented over time as a sinusoidal curve with varying amplitudes, similar to that shown in plot 520.

[0130]

[0130] The excitation voltage 515 can be an alternating current at a predetermined drive frequency. The particular frequency selected can depend for example upon the particular target sought to be detected, the medium of the test sample, the chemical makeup of the amplification process constituents, the temperature of the amplification process, and / or the excitation voltage. In some embodiments, the excitation drive frequency can be between 1 kHz and 10 kHz at as low an excitation voltage as possible. As one example, in tests performed to identify a target of H. influenzae (106copies / reaction) spiked into 5% whole blood, excitation sensor drive frequency was varied from 100 Hz to 100,000 Hz at 0.15 Volts. These tests revealed that the desired “signal cliff,” an artifact in a portion of the signal indicative of a positive test sample described in more detail below, becomes more easily detectable below 100 Hz and is most easily detectable between 1 kHz and 10 kHz. Further, with frequencies in the range between 1 kHz and 10 kHz, the signal cliff advantageously could be identified before 12 minutes of test time had elapsed. Beneficially, faster identification of the signal cliff can result in shorter test times, in turn resulting in quicker provision of test results and the ability to perform more tests per day. At frequencies lower than 1 kHz, the reactance component of the signal (in which the signal cliff may be found in a positive sample) decreased monotonically. The sensor drive frequency can be similarly fine-tuned for other tests to optimize performance, that is, to optimize the detectability of a signal cliff. Detectability of a signal cliff refers to the ability to consistently differentiate between a positive sample and a negative sample.

[0131]

[0131] FIG. 5B depicts an example plot 525 showing an impedance signal 530 that can be extracted from the raw signal 520 provided by the sensing electrode 51 OB. The impedance signal 530 represents the electrical impedance Z of the test well over time. The impedance Z can be represented by a Cartesian complex number equation as follows:

[0132] Z = R +jX where R represents the resistance of the test well and is the real part of the above equation and the X represents the reactance of the test well and is the imaginary part of the above equation (denoted by j). Thus, the impedance of the test well can be parsed into two components, the resistance R and the reactance X.

[0133]

[0132] Initially, the value of the resistance R can be determined by taking a baseline measurement of the test well prior to or at the outset of the amplification process. Although the resistance of the test fluid can drift away from this baseline value throughout the duration of the test, the current sensed by the sensing electrode 51 OB due to the resistance of the test fluid can be in phase with the signal provided through the excitation electrode 510A. Thus, changes or drift in the resistance can be identified by values of the in-phase component of the signal 520 over time. The reactance can arise from the effect of inductance in the test fluid, capacitance in the test fluid, or both; this effect can cause the fluid to retain current (e.g., electrons provided by excitation electrode 510A) temporarily. After some time, this retained current flows out of the test fluid into the sensing electrode 51 OB. Due to this delay, the current sensed by the sensing electrode 51 OB due to the reactance of the test fluid can be out of phase with the current sensed from the resistance of the test fluid. Thus, values of the reactance of the test fluid can be identified by values of the out of phase component of the signal 520 over time. The reactance can fluctuate throughout the duration of the test based on changes to the chemical constituents of the test fluid due to the amplification process. The signal cliff (e.g., a rise or drop in the reactance at or greater than a threshold rate or magnitude and / or during a predetermined window of time) indicative of a positive sample can be found in the reactance X.

[0134]

[0133] During a test, the excitation electrode 510A can be sinusoidally excited with some amplitude and voltage. The excitation electrode 510A is in series with the test liquid in the well, which can be considered as a resistor R. The resistor (e.g., the test fluid) and electrode form a voltage divider, which has a voltage determined by the ratio of the resistor and electrode chemistry / impedances. The resulting voltage waveform sensed at the sensing electrode 51 OB represents the complex impedance signal 530. In some embodiments, a curve such as the impedance signal 530 may not be generated, but rather the raw sensed signal 520 can be parsed into its resistance and reactance components as described herein. The impedance signal 530 is provided as an example representation of a combined curve representing both the resistance of the test fluid and the reactance of the test fluid over time. The complex impedance signal 530 can be interpreted as a quadrature-modulated waveform (e.g., a combination of an in-phase waveform resulting from the resistance of the test fluid and an out-of-phase waveform resulting from the reactance of the test fluid), where the in- phase and out-of-phase components change on a timescale much greater than the modulation frequency. The in-phase waveform is in-phase with the composite waveform of the complex impedance. Some implementations can use a synchronous detector, for example having multipliers and low pass filters implemented in a field programmable gate array (FPGA), to extract the in-phase and out-of-phase components from the raw signal 520 and compute their amplitude and phase.

[0135]

[0134] In order to parse the impedance signal 530 (or the raw sensed signal 520) into its constituent resistance and reactance components, the voltage waveform 520 at the sensing electrode 510B is sampled faster than its Nyquist frequency (e.g., two times the highest frequency of the excitation voltage) and then decomposed into an in-phase component (resistance) and an out-of-phase component (reactance). The in-phase and out-of-phase voltage components can be computed using the known series resistance (e.g., the value of R) to calculate the real component of the impedance (the resistance) and the imaginary component of the impedance (the reactance).

[0136]

[0135] FIG. 5C depicts a plot 541 of the resistance 540A and reactance components 540B over time (t = 3 minutes to t = 45 minutes) extracted from a raw signal 520 generated based on an example positive test. As illustrated, the signal cliff 545 represents a change AR in the reactance 540B during a particular window of time Tw. The signal cliff 545 indicates a positive sample. At times occurring prior to the signal cliff 545, the reactance curve 540B is relatively flat or stable, and again after the signal cliff 545 the reactance curve 540B is relatively flat or stable. Thus, in this embodiment the signal cliff 545 for the particular test parameters represented by the plot 541 occurs as a drop of AR in the expected region 535.

[0137]

[0136] The magnitude of the change AR in the reactance that corresponds to a positive sample signal cliff 545, as well as the position and / or duration of the particular window of time Tw at which the signal cliff 545 is expected to occur, can vary depending on a number of parameters of the test. These parameters include the particular target of the test (e.g., the rate at which that target amplifies), the frequency of the excitation voltage, the configuration of the excitation and sensor electrodes (e.g., their individual shapes and dimensions, the gap separating the electrodes, and the material of the electrodes), the sampling rate, the quantity of amplification agents provided at the start of the test, the temperature of the amplification process, and the amount of target present in the sample. In some embodiments, the expected characteristics of a signal cliff of a positive sample, predetermined for example through experimentation, can be used for differentiating between positive samples and negative samples. In some embodiments, the expected characteristics of a signal cliff can be used for determining the severity or progress of a medical condition, for example via correlations between particular signal cliff characteristics and particular initial quantities of the target in the sample. The predetermined expected characteristics can be provided to, stored by, and then accessed during test result determination by a reader device configured to receive signals from the sensing electrode(s) of a test cartridge.

[0138]

[0137] For a given test, the expected magnitude of the change AR in the reactance and the expected window of time Tw of a signal cliff 545 for a positive sample can be determined experimentally based on monitoring and analyzing the reactance curves generated by positive control samples (and optionally negative control samples). In some embodiments, the test parameters influencing the signal cliff can be varied and fine-tuned to identify the parameters that correspond to an accurately distinguishable signal cliff A reader and cartridge as described herein can be configured to match the tested configuration and provided with expected signal cliff characteristics for that test.

[0138] For example, in a set of experimental tests for H. influenzae, the test fluid initially included amplification primers and 1,000,000 added target copies, the excitation voltage was 200 mV P2P, the test parameters included a 10 kHz sweep start and a 10 MHz sweep stop for the frequency of the excitation current, and close and far electrode gaps were configured at 2.55 mm and 5 mm respectively. The amplification temperature was set to 65.5 degrees Celsius, and the two electrode setups (one for each of the close and far gaps) included platinum electrodes. At low frequencies (10 kHz-lOOkHz), detectable signal cliffs were identified beginning around 23 minutes into amplification around 10 kHz and around 30 minutes around 100 kHz using the 5 mm gap electrode configuration, with the magnitude of change in reactance being around 3.5-4 Ohms at 10 kHz and dropping to around 3.25-3.5 Ohms at 100 kHz. At low frequencies (10 kHz-lOOkHz), detectable signal cliffs were identified beginning around 25 minutes into amplification around 10 kHz and around 30 minutes around 100 kHz using the 2.5 mm gap electrode configuration, with the magnitude of change in reactance being around 3.5-4 Ohms. At higher frequencies, the drop in reactance of the signal cliff decreased, and the time at which these smaller signal cliffs were identified was shifted to later in the amplification process. Accordingly, in this example a test well in a test cartridge may be configured with the 5 mm gap electrodes and a reader device may be configured to provide 10 kHz excitation current to the test cartridge during amplification. The reader device can be provided with instructions to provide this current and monitor the resulting reactance of the test well throughout amplification or for a window of time around the expected signal cliff time (here, 23 minutes), for example between 20 and 35 minutes. The reader device can also be provided with instructions to identify a positive sample based on the reactance exhibiting around a 3.5-4 Ohm change around 23 minutes into amplification, or within the window of time around the expected signal cliff time.

[0139]

[0139] Once identified, the values for AR and Tw can be provided to reader devices for use in distinguishing between positive and negative samples for that particular test. In some examples, such devices can determine whether the reactance curve 540B has the required value and / or slope at the identified window of time Tw to correspond to the signal cliff In other embodiments, the reader device can analyze the shape of the reactance curve over time to determine whether it contains a signal cliff In some embodiments, a reader can modify its testing procedures based on the identified window of time Tw at which the signal cliff 545 is expected to occur, for example by only providing the excitation voltage and monitoring the resultant signal within this window, advantageously conserving power and processing resources compared to continuous monitoring during an entire test time.

[0140]

[0140] FIG. 5D depicts a plot 551 of the resistance and reactance components extracted from the raw sensor data of a sensing electrode 51 OB during example tests of positive and negative controls. Specifically, the plot 551 shows a curve 550A of the resistance of the positive sample, a curve 55 OB of the reactance of the positive sample, a curve 550C of the resistance of the positive sample, and a curve 550D of the reactance of the positive sample over the 35 minute duration of the test. As shown by FIG. 5D, the positive sample signal cliff occurs around 17 minutes into the test, with a relatively flat and stable reactance curve 550B leading up to the signal cliff In contrast, at this same time the negative sample reactance curve 550D exhibits no signal cliff, but rather maintains a quadratic curvature from around t = 8 minutes through the end of the test.

[0141]

[0141] FIG. 5E depicts a plot 561 of the resistance 560A and reactance components 560B over time (t = 0 minutes to t = 60 minutes since the start of amplification) extracted from a raw signal 520 generated based on an example positive test. As illustrated, the signal cliff 565 represents a change AR in the reactance 560B during a particular window of time Tw. The signal cliff 565 indicates a positive sample. At times occurring prior to the signal cliff 565, the reactance curve 560B is relatively flat or stable, and again after the signal cliff 565 the reactance curve 560B is relatively flat or stable with slight concavity. The signal cliff 565 for the particular test parameters represented by the plot 561 occurs as a peak, spike, or bell curve in the expected region 535, during which the reactance values rise and fall by the AR value in an approximately parabolic curve. As described herein, varying of certain test parameters (e.g., test well configuration, chemistry and initial quantity of amplification constituents, target, and excitation current characteristics) can vary the geometry of the signal cliff yielded from a positive sample. Thus, in some embodiments the geometry of a “signal cliff’ in the reactance values vs time curve can vary from test to test, though for a particular test the curve geometry and / or timing signal cliff remains consistent within reactance change and / or timing parameters across positive samples for that test.

[0142]

[0142] FIG. 6 depicts a schematic block diagram of an example reader device 600 that can be used with the cartridges described herein, for example the cartridge 100. The reader device 600 includes a memory 605, processor 610, communications module 615, heater 625, electrode interface 630, voltage source 635, and a cavity 660 into which a cartridge can be inserted. The reader device 600 may further include a status indicator 640. The reader device 600 is in communication with a user interface 620, which may include a user interface of a remote computing device such as a smartphone, tablet, or other device having a testing control application executing thereon.

[0143]

[0143] When test cartridge 100 is inserted into the cavity 660 of the reader device 600, the electrode interface 214 of the cartridge couples with the electrode interface 630 of the reader device 600. This can allow the reader device 600 to detect that a cartridge is inserted, for example by testing whether a communication path is established. In some embodiments, an optional power cartridge may activate a power supply circuit of the reader device 600 when the electrode interface 214 of the cartridge couples with the electrode interface 630 of the reader device 600. Further, such communications can enable the reader device 600 to identify a particular inserted test cartridge 100 and access corresponding testing protocols. Testing protocols can include the duration of the test, the temperature of the test, the characteristics of a positive sample impedance curve, and the information to output to the user based on various determined test results. In other embodiments, the reader device 600 can receive an indication via user interface 620 that a cartridge is inserted (e.g., by a user inputting a “begin testing” command and optionally a test cartridge identifier).

[0144]

[0144] The memory 605 includes one or more physical electronic storage devices configured for storing computer-executable instructions for controlling operations of the reader device 600 and data generated during use of the reader device 600. For example, the memory 605 can receive and store data from sensing electrodes coupled to the electrode interface 630.

[0145]

[0145] The processor 610 includes one or more hardware processors that execute the computer-executable instructions to control operations of the reader device 600 during a test, for example by controlling the heater 625, controlling the communications module 615 to interact with the user interface 620, and activating the voltage source 635. One example of testing operations is described with respect to FIG. 7A below. The processor 610 can be also be configured by the instructions to determine test results based on data received from the excitation electrodes of an inserted test cartridge, for example by performing the process of FIG. 7B described below.

[0146]

[0146] The communications module 615 includes network-enabled hardware components, for example wired or wireless networking components, for providing networked communications between the reader device 600 and remote computing devices. Suitable networking components include WiFi, Bluetooth, cellular modems, Ethernet ports, or USB ports, and the like. Beneficially, networking capabilities can enable the reader device 600 to interact with and be controlled by remote computing devices such as one or more additional handheld computing devices (e.g., smartphones, tablets, etc.). In some embodiments, remote devices may be in communication additional remote computing systems such as hospital information systems and / or laboratory information systems that store electronic medical records, national health agency databases, and the computing devices of clinicians or other designated personnel. In addition, the networking capabilities can enable the reader device 600 to receive information over the network from remote computing devices, for example updated signal cliff parameters for existing test, new signal cliff parameters for new tests, and updated or new testing protocols.

[0147]

[0147] The user interface 620 can be implemented within a remote device connected to the communications module 615 via WiFi, or Bluetooth, or the like. The remote device may have a testing control application installed thereon to provide a testing system user interface, for providing control options and / or presenting test results and other test information to users, on a display of the remote device.

[0148]

[0148] The heater 625 can be positioned adjacent to the cavity 660 for heating an inserted cartridge to the desired temperature for an amplification process. Though depicted on a single side of the cavity 660, in some embodiments the heater 625 can surround the cavity.

[0149]

[0149] As described herein, the voltage source 635 can provide an excitation signal at a predetermined voltage and frequency to the excitation electrode of an inserted test cartridge.

[0150]

[0150] The status indicator 640 may include any suitable notification device, such as one or more lights, sound generators, or the like.

[0151] FIG. 7A depicts a flowchart of an example process 700 for operating a reader device during a test as described herein. The process 700 can be performed by the reader device 600 described above.

[0151]

[0152] At block 705, the reader device 600 can detect that a power cartridge has been removed from the reader device 600. In some embodiments, the detection of block 705 can occur based on the disconnection of a signal path between the electrode interface 630 of the reader device 600 and one or more contact pads of the power cartridge.

[0152]

[0153] At block 710, the reader device 600 automatically powers on in response to detecting the removal of the power cartridge at block 705. In some embodiments, the reader device may transmit a notification to a user interface 620 device and / or illuminate one or more status lights of a status indicator 640 to indicate that the reader device 600 is powered on and ready to receive an assay cartridge 100.

[0153]

[0154] At block 715, the reader device 600 can detect that an assay cartridge 100, 200 has been inserted, for example in response to user input or in response to establishing a signal path with the inserted cartridge. In some embodiments, the cartridge 100, 200 can include an information element that identifies the particular test(s) to be performed to the reader device 600 and optionally includes test protocol information.

[0154]

[0155] At block 720, the reader device 600 can heat the cartridge 100, 200 to a specified temperature for amplification. For example, the temperature can be provided by information stored on the cartridge 100, 200 or accessed in the internal memory of the reader device 600 in response to identification of the cartridge 100, 200.

[0155]

[0156] At decision block 725, the reader device 600 can determine whether the test is still within its specified test duration. For example, where the expected window of time in which a signal cliff should appear in a positive sample is known, the duration of the test may end at or some predetermined period of time after the end of the window. If so, the process 700 transitions to optional decision block 730 or, in embodiments omitting block 730, to block 735.

[0156]

[0157] At optional decision block 730, the reader device 600 determines whether to monitor the test well amplification by logging data from the test well sensing electrode. For example, the reader 600 may be provided with instructions to only monitor the impedance of the test well during a particular window or windows of a test. If the reader device 600 determines not to monitor the test well amplification, the process 700 loops back to decision block 725.

[0157]

[0158] If the reader device 600 determines to monitor the test well amplification, the process 700 transitions to block 735. At block 735, the reader device 600 provides an excitation signal to the excitation electrode of the test well(s) of the inserted cartridge. As described above, this can be an alternating current at a particular frequency and voltage.

[0158]

[0159] At block 740, the reader device 600 detects and logs data from the sensing electrode of the test well(s) of the inserted cartridge. In some embodiments, this data can be stored for later analysis, for example after completion of the test. In some embodiments, the reader device 600 can analyze this data in real time (e.g., as the test is still occurring) and may stop the test once a positive sample signal cliff is identified.

[0159]

[0160] When the reader device 600 determines at block 725 that the test is not still within its specified duration, the process 700 moves to block 745 to analyze the test data and output the test result. The test result can include an indication that the sample tested positive or negative for the target or can more specifically indicate an estimated quantity of the target in the tested sample. Following the conclusion of the test, further tests may be performed by returning to block 715 for a new assay cartridge. Alternatively, the reader device 600 may detect insertion of a power cartridge and power off in response.

[0160]

[0161] FIG. 7B depicts a flowchart of an example process 750 for analyzing test data to detect a target as described herein that can be performed by the reader device 600 as block 745 of FIG. 7A.

[0161]

[0162] At block 755, the reader device 600 can access logged signal data received from the electrode of a well.

[0162]

[0163] At block 760, the reader device 600 can decompose the signal into resistance and reactance components across some or all of the different time points of the test. For example, as described above, at each time point the reader device 600 can determine in phase and out of phase components of the raw sampled voltage waveform and can then deconvolute these components using known series resistance of the electrode circuit to calculate the in- phase (resistance) and out-of-phase (reactance) portions of the impedance of the test well.

[0164] At block 765, the reader device 600 can generate a curve of the reactance values over time. Also, at block 765, the reader device 600 can optionally generate a curve of the resistance values over time.

[0163]

[0165] At block 770, the reader device 600 can analyze the reactance curve to identify a signal change indicative of a positive test. As described above with respect to the signal cliff of FIG. 5C, the reader device 600 can look for greater than a threshold change in reactance, can look for such a change within a predetermined window of time, can analyze the slope of the reactance curve at a predetermined time, or can analyze the overall shape of the reactance curve in order to determine whether a signal cliff (e.g., a rise or drop in the signal preceded and followed by relatively more stable values) is present.

[0164]

[0166] At decision block 775, based on the analysis performed at block 770, the reader device 600 can determine whether the sought-after signal change was identified in the reactance curve. If so, the process 750 transitions to block 780 to output an indication of a positive test result to the user. If not, the process 750 transitions to block 785 to output an indication of a negative test result to the user. The result can be output locally, for example on the display of the device, or output over a network to a designated remote computing device.

[0165]

[0167] The various components of the handheld detection systems (for example, the analyzers or reader device 300 and cartridge 100, described herein) may be integrated with an external computing device. The external computing device, for example a mobile phone, a computer, a tablet, a laptop, or similar device, may communicate with the reader device 300 using a communication interface. The external computing device may comprise a testing monitoring and / or control application installed thereon to provide a user options and abilities to monitor and / or control one or more of the cartridge or the reader device.

[0166]

[0168] In some embodiments, the user interface works with the reader device (for example, the reader devices 300) to determine whether a subject is suffering from a particular illness. For example, the reader device may perform a test using the cartridge 100 and determine that a particular target agent is present in a sample from the subject. However, merely the presence and / or amount of the target agent may be insufficient to determine that the subject is suffering from an illness or ailment. For example, if the subject is merely a carrier of an illness, the subject test results may indicate presence and / or amount of the virus but the subject may not be suffering from an associated illness. Thus, the user interface may receive symptoms that the subject is experiencing and combine these symptoms with the test results from the reader device to determine whether the subject is suffering from an illness or ailment corresponding to the target agent. For example, if the test result from the reader device indicates the presence and / or amount of the avian flu virus in the subject’s mucus but the subject is not experiencing any symptoms, then the user interface may determine that the subject is not suffering from the influenza virus but is instead a carrier for the virus. On the other hand, when the test result indicates the presence and / or amount of the avian flu virus and the subject’s symptoms are known to coincide with the avian flu virus in a subject suffering from the influenza illness, then the user interface may determine that the subject is infected with the influenza illness. Such a determination may be based on a threshold number of symptoms or specific being met while the target agent is detected (for example, one or two symptoms being suffered while the avian flu virus is present). The symptoms may be weighted differently depending on the target agent in the test and / or the corresponding illness. For example, when the target agent is avian flu virus, then symptoms such as fever, chills, muscle aches may be more highly weighted than symptoms such as headache or cough. Thus, symptoms associated with a corresponding illness for the target agent may have higher weights and be more indicative that the subject carrying the virus is suffering the corresponding illness. In some embodiments, the threshold number of symptoms, weighting of symptoms, or specific symptoms to be met to determine that the subject is ill is determined based on one or more metrics. A standard setting or national organization, such as the Center for Disease Control (CDC) or similar organization or entity, may establish the one or more metrics. Thus, the user interface may use the test results from the reader device and the symptom information provided by the user to determine whether the subject is (1) ill or sick or (2) a carrier for the target agent. In some embodiments, the user interface offloads one or more of the determinations described herein to one or more external systems with which the user interface (and the corresponding external computing device) interacts.

[0167]

[0169] In some embodiments, the user interface may generate a score or similar indicator to indicate a probability that the subject is ill or sick. A higher score may correspond to a higher probability that the subject is ill while a lower score may correspond to a higher probability that the subject is not ill. For example, the user interface may use details from the test results from the reader device along with the user provided symptom information to generate the score indicating the probability of sickness of the subject.

[0168]

[0170] The reader device may provide test results that include a range of values, where the range of values correspond to a range of possible detection levels of the target agent. For example, the test results may include one or more of a likelihood that the mucus sample included the target agent (for example, between 0 and 100% probability) and a quantity of the target agent determined to have been included in the mucus sample. The probability that the mucus sample included the target agent may be associated with the quantity of the target agent determined to have been included in the mucus. The range of possible detection levels may refer to different likelihoods or probabilities that the subject has a particular virus or infection. For example, if the reader device provides test results indicating that the subject does have the virus or infection with 100% certainty or that a quantity of the target agent above a first threshold was present, then the score (or probability) may be assigned a minimum value, for example 50 out of a 0 to 100 range. Then, if the subject is experiencing any symptoms associated with the illness associated with the virus or infection, then the value of the score (indicating that the subject is suffering from the corresponding illness) may be increased. For example, for each symptom that the subject experiences that is associated with the illness, the value may increase by a threshold value (for example, 10 points). Thus, the combination of the target agent detection and the symptoms can increase or decrease the score or probability of the subject being sick. Alternatively, if the reader device indicates that the subject has the virus or infection with 50% certainty (or that a quantity of the target agent below the first threshold but above a second threshold was present), then the score may be assigned a different minimum value, for example 25 out of the 0 to 100 range. Accordingly, when the test results indicate a lower probability of the presence of the target agent, more symptoms suffered by the subject are needed to increase the score to the same value as compared to when the test results indicate a high probability of sickness. If the reader device indicates that the subject does not have the virus or infection (or that a quantity of the target agent below the second threshold was present), then the score may be assigned a zero value, for example 0 out of the 0 to 100 range. When the score starts at a zero-value based on the test results, no quantity of symptoms may be sufficient to raise the score because the virus or target agent is not present to make the subject ill. Thus, the user interface and the reader device may together generate the score that represents the probability of the subject being sick based on the test results of the target agent and the symptoms suffered by the subject.

[0169]

[0171] In some embodiments, the user interface may review the test results and the provided symptoms to determine whether the subject is suffering from a particular illness not associated with the target agent for which the test was run. As such, the user interface may use knowledge (for example, databases) of similarities and differences in symptoms of different illnesses but differences in test results and so forth in illness determinations.

[0170]

[0172] In some embodiments, the user interface generates a score value that indicates a probability that the subject is sick or ill, based on the test results and the symptom information. The user interface may further aggregate external information into the score value, regardless of what the test results and symptoms by themselves would otherwise indicate. The external information may include one or more of test results from other subjects that used the same reader device, test results from other reader devices that interfaced with the user interface and the external computing device, test results and other diagnostic information for subjects in a clinic or a geographic region, and so forth. For example, the user interface may use test results and / or symptom information from other subjects tested by the reader device to inform further the score values. For example, many other subjects tested with the reader device coupled to the user interface have positive test results for the same target agent and report one or more similar symptoms. The user interface may use this information to increase a score or probability that the subject is ill or sick, even if the subject’s test results and / or symptoms may alone not indicate that the subject is ill. Similarly, the test results and the symptom information for the subject may indicate that the subject is ill but other subjects having similar test results and symptoms report that they are not ill. The user interface then may decrease the score or probability that the subject is ill, regardless of what the subject’s test results and / or symptom information otherwise indicate. If the test results and the symptoms for the subject are different from those of other subjects that report as not being sick (for example, by a threshold amount), then the user interface may increase the score or probability that the subject is sick. On the other hand, if the test results and symptoms are different from those of other subjects that report as being sick (for example, by the threshold amount), then the user interface may decrease the score or probability that the subject is sick, regardless of the test results and symptoms.

[0171]

[0173] In some embodiments, the user interface performs one or more actions based on the score or probability values. The user interface may communicate information to one or more of the user, the subject, attending medical staff, the CDC, or similar entities. Alternatively, or additionally, the user interface may generate an alert to the one or more of the user, the subject, attending medical personnel, the CDC, or similar entities. The alert may comprise one or more of a phone call, a text message, an e-mail message, a push message, an audio message, a flashing indicator, or audible indicator, or any other communication used to communicate information. For example, the user interface may automatically generate and transmit an alert to the user based on the results of the test or the indicated symptoms, a combination of the two, or the test results or symptoms in aggregate with information from other subjects. For example, if the test results and the indicated symptoms suggest that the subject is suffering from a rare illness, then the user interface may generate and communicate an alert to the subject or user suggesting a follow-up visit to a specialist. Similarly, if the test results and the indicated symptoms suggest that the subject is suffering from highly contagious illness, then the user interface may generate and communicate the alert to the subject or user requesting that the subject restrict interaction with others to minimize risk of communication of the illness to others. If the test results and the indicated symptoms suggest that the subject is suffering from a highly contagious, rare, and difficult to treat illness, then the user interface may generate and communicate the alert to the subject or user but also to local, regional, or national medical staff or disease monitoring agencies. As such, the reader device, cartridge, and user interface on the external computing device may generate information used to help detect an outbreak of a virus or disease.

[0172]

[0174] For example, the user interface and the external computing device may communicate and / or interact with a system that tracks illnesses over a geographic area, for example a city, county, state, or nation, or a specific portion of the population. Thus, the user interface and the external computing device may enable tracking of diseases and / or infections by the system for a number of subjects in various geographic areas. In some embodiments, the system performs tracking based on one or both of the test results and the identified symptoms. In some embodiments, the system performs the tracking based on the scores or probabilities generated by the user interface. The system may aggregate and use the information from multiple user interfaces and external computing devices to generate a geographic heat map. The heat map may show levels of one or more illnesses and / or corresponding rates of infection or healing as different colors or levels on the map, where different colors correspond to different levels (for example, numbers) of reported injections or rates of infection. Thus, the heat map may visually show how numbers of ill subjects vary in the geographic area. In some embodiments, the system may use such heat maps to identify pockets of specific illnesses or infections and / or an epidemic that is occurring based on aggregated test results and symptoms provided from multiple user interfaces and corresponding external computing devices. The heat map may show different quantities of illnesses or different rates of illness detection in different colors. The heat map may enable an entity to review quickly the geographic area to identify relational information (for example, information for portions of the geographic area relative to one another) and specific information (for example, detailed information for individual portions of the geographic area).

[0173]

[0175] In some embodiments, the user interface and the external computing device may be integrated with a system (for example, the system that generates the heat map) used to track and / or determine need for vaccines or other medications. The system may receive test results, symptoms, and / or scores from a number of user interfaces and corresponding external computing devices. Such aggregation of results may allow for early detection of outbreaks. Such automatic detection of supply need and request for additional supplies may improve response times in times of outbreaks and help prevent and / or reduce the spread of communicable diseases.

[0174]

[0176] Similarly, such aggregate tracking of test results, symptoms, and / or scores may allow the system to provide recommendations. Such recommendations may include increased education, advertisement, and so forth.

[0175]

[0177] In some embodiments, the user interface may also generate an indicator to indicate whether the subject is becoming sicker or healthier. For example, if the user performs multiple tests for the subject, then the user interface may determine that different combinations of symptoms suffered by the subject or different quantities of the target agent in the test sample indicate whether the subject is sicker than a previous test or healthier than a previous test.

[0176]

[0178] In some embodiments, the indicator may comprise an arrow or representation of a face, or similar indicator.

[0177] Overview of Example Devices

[0178]

[0179] Some embodiments of the methods, systems and compositions provided herein include devices comprising an excitation electrode and a sensor electrode. In some embodiments, the excitation electrode and the sensor electrode measure electrical properties of a sample. In some embodiments, the electrical properties comprise complex admittance, impedance, conductivity, resistivity, resistance, and / or a dielectric constant.

[0179]

[0180] In some embodiments, the electrical properties are measured on a sample having electrical properties that do not change during the measurement. In some embodiments, the electrical properties are measured on a sample having dynamic electrical properties. In some such embodiments, the dynamic electrical properties are measured in real-time.

[0180]

[0181] In some embodiments, an excitation signal is applied to the excitation electrode. The excitation signal can include direct current or voltage, and / or alternating current or voltage. In some embodiments, the excitation signal is capacitively coupled to / through a sample. In some embodiments, the excitation electrode and / or the sensor electrode is passivated to prevent direct contact between the sample and the electrode.

[0181]

[0182] In some embodiments, parameters are optimized for the electric properties of a sample. In some such embodiments, parameters can include the applied voltage, applied frequency, and / or electrode configuration with respect to the sample volume size and / or geometry.

[0182]

[0183] In some embodiments, the voltage and the frequency of the excitation voltage may be fixed or varied during the measurement. For example, measurement may involve sweeping voltages and frequencies during detection, or selecting a specific voltage and frequency which may be optimized for each sample. In some embodiments, the excitation voltage induces a current on the signal electrode that can vary with the admittance of the device and / or sample characteristics.

[0184] In some embodiments, the detection parameters are optimized by modeling the admittance, device and sample by the lumped-parameter equivalent circuit consisting of electrode-sample coupling impedances, sample impedance, and inter-electrode parasitic impedance. Parameters of the lumped-parameter equivalent circuit is determined by measuring the admittance of the electrode-sample system at one or many excitation frequencies for a device. In some embodiments, the complex (number having both real and imaginary components) admittance of the electrode-sample system is measured using both magnitude- and phase-sensitive detection techniques. In some embodiments, the detection parameters are optimized by determining the frequencies corresponding to the transitions between the frequency regions by measuring the admittance across a wide range of frequencies. In some embodiments, the detection parameters are optimized by determining the frequencies corresponding to the transitions between the frequency regions by computing from the values given lumped-parameter model.

[0183]

[0185] In some embodiments, the admittance of a capacitively-coupled electrodesample system comprises three frequency regions: a low frequency region dominated by the electrode-sample coupling impedance, a mid-frequency region dominated by the sample impedance, and a high frequency region dominated by parasitic inter-electrode impedance. The admittance in the electrode-sample coupling region is capacitive in nature and is characterized by a magnitude that increases linearly with frequency, whose phase is ninety degrees. The admittance in the sample region is conductive in nature and is characterized by an admittance that does not vary significantly with respect to frequency, whose phase is approximately zero degrees. The admittance inter-electrode region is capacitive in nature and is characterized by a magnitude that increases linearly with frequency and a phase of ninety degrees.

[0184]

[0186] In some embodiments, an induced current at the pick-up electrode is related to the excitation voltage and complex admittance by the relation: current = (complex admittance) X (voltage)

[0185]

[0187] In some embodiments, the device measures both the excitation voltage magnitude and induced current magnitude to determine the magnitude of the complex admittance. In some embodiments, the device is calibrated to known excitation voltages and measure the magnitude of the induced current. In order to determine the phase of complex admittance, the device may measure the relative phase difference between the excitation voltage and the induced current.

[0186]

[0188] In some embodiments, the magnitude and phase are measured directly.

[0187]

[0189] In some embodiments, the magnitude and phase are measured indirectly e.g., by using both synchronous and asynchronous detection. The synchronous detector gives the in-phase component of the induced current. The asynchronous detector gives the quadrature component of the induced current. Both components can be combined to determine the complex admittance.

[0188]

[0190] In some embodiments, the electrodes are not passivated.

[0189]

[0191] In some embodiments, the excitation and / or detection electrodes are passivated. The excitation and / or detection electrodes may be passivated to prevent e.g., undesirable adhesion, fouling, adsorption or other detrimental physical interactions between the electrode with the sample or components therein. In some embodiments, the passivation layer comprises a dielectric material. In some embodiments, passivation enables efficient capacitive coupling from the electrodes to the sample. The efficiency of the coupling is determined by measuring the characteristics of the electrode / sample system, for example, which may include: the dielectric properties of the passivation layer, the thickness of the passivation layer, the area of the passivation / sample interface, the passivation surface roughness, the electric double layer at the sample / passivation interface, temperature, applied voltage and applied frequency, the electrical properties of the sample, the electric and / or chemical properties of the electrode materials.

[0190]

[0192] In some embodiments, the electrode configuration and fabrication are optimized to mitigate undesirable parasitic coupling between electrodes. This may be accomplished through electric field shielding, the use of a varying dielectric constant electrode substrate, layout optimization, and / or grounding layers.

[0191]

[0193] In some embodiments, the electrode configuration and fabrication are not optimized to mitigate undesirable parasitic coupling between electrodes.

[0192] Implementing Systems and Terminology

[0193]

[0194] Implementations disclosed herein provide systems, methods and apparatus for detection of the presence and / or quantity of a target analyte. One skilled in the art will recognize that these embodiments may be implemented in hardware or a combination of hardware and software and / or firmware.

[0194]

[0195] The signal processing and reader device control functions described herein may be stored as one or more instructions on a processor-readable or computer-readable medium. The term “computer-readable medium” refers to any available medium that can be accessed by a computer or processor. By way of example, and not limitation, such a medium may comprise RAM, ROM, EEPROM, flash memory, CD-ROM or other optical disk storage, magnetic disk storage or other magnetic storage devices, or any other medium that can be used to store desired program code in the form of instructions or data structures and that can be accessed by a computer. It should be noted that a computer-readable medium may be tangible and non-transitory. The term “computer-program product” refers to a computing device or processor in combination with code or instructions (e.g., a “program”) that may be executed, processed or computed by the computing device or processor. As used herein, the term “code” may refer to software, instructions, code or data that is / are executable by a computing device or processor.

[0195]

[0196] The various illustrative logical blocks and modules described in connection with the embodiments disclosed herein can be implemented or performed by a machine, such as a general purpose processor, a digital signal processor (DSP), an application specific integrated circuit (ASIC), a field programmable gate array (FPGA) or other programmable logic device, discrete gate or transistor logic, discrete hardware components, or any combination thereof designed to perform the functions described herein. A general purpose processor can be a microprocessor, but in the alternative, the processor can be a controller, microcontroller, combinations of the same, or the like. A processor can also be implemented as a combination of computing devices, e.g., a combination of a DSP and a microprocessor, a plurality of microprocessors, one or more microprocessors in conjunction with a DSP core, or any other such configuration. Although described herein primarily with respect to digital technology, a processor may also include primarily analog components. For example, any of the signal processing algorithms described herein may be implemented in analog circuitry. A computing environment can include any type of computer system, including, but not limited to, a computer system based on a microprocessor, a mainframe computer, a digital signal processor, a portable computing device, a personal organizer, a device controller, and a computational engine within an appliance, to name a few.

[0196] Additional Definitions

[0197]

[0197] Disclosed herein are methods of detecting the presence and / or amount of a nucleic acid in a biological sample. In some embodiments, the biological sample is obtained from a subject, preferably a human or other animal, a plant, a food, soil, or a surface, or any combination thereof. In some embodiments, the biological sample is obtained by swabbing. In some embodiments, the biological sample is or is derived from saliva, nasal wash, nasal swab, nasal nasopharyngeal swab, oropharyngeal swab, mucus, lavage fluid, blood, plasma, urine, stool, serum, cerebral spinal fluid, or any material comprising a pathogen such as one or more of a microbe, virus, bacteria, mold, or fungus. In some embodiments, the subject is a human. In some embodiments, the subject is a mammal. The methods comprise contacting the biological sample with an assay cartridge or a detection system, amplifying the nucleic acid by loop-mediated isothermal amplification (LAMP) with a primer set, measuring or analyzing a modulation of an electrical signal for the duration of the amplification with the primer set using the detection system, thereby detecting successful amplification of the nucleic acid with the primer set, and determining the presence and / or amount of the nucleic acid in the biological sample. In some embodiments, the nucleic acid is a nucleic acid from a pathogen. In some embodiments, the primer set comprises, consists essentially of, or consists of one or more F3 primers, one or more B3 primers, one or more FIP primers, and one or more BIP primers. In some embodiments, the primer set comprises, consists essentially of, or consists of one or more F3 primers, one or more B3 primers, one or more LF primers, one or more LB primers, one or more FIP primers, or one or more BIP primers, or any combination thereof. In some embodiments, the primer set comprises, consists essentially of, or consist of one or more F3 primers, one or more B3 primers, one or more LF primers, one or more LB primers, one or more FIP primers, and one or more BIP primers. Each of the F3 primers, B3 primers, LF primers, LB primers, FIP primers and BIP primers are used for LAMP. The FIP primers and BIP primers are required for the LAMP process, whereas the F3 primers, B3 primers, LF primers and / or LB primers strongly enhance the amplification but optionally may be excluded for high concentration targets. In some embodiments, each of the one or more F3 primers, one or more B3 primers, one or more LF primers, one or more LB primers, one or more FIP primers, or one or more BIP primers comprise 1, 2 ,3, 4, 5, 6, 7, 8, 9, or 10 primers. In some embodiments, the pathogen comprises more than one population obtained from different sources that may exhibit variations in their genes or genome regions (e.g., serotypes, strains, mutants, isolates, species, variants, types, subtypes, or clones) and the inclusion of more than one primer may increase amplification of the pathogen comprising more than one population. In some embodiments, the primer set is specific for a gene or a genome region of the pathogen. In some embodiments, the gene or the genome region is associated with the pathogenicity of the pathogen. In some embodiments, the nucleic acid is DNA, RNA, both, or a fragment or hybrid thereof. In some embodiments, where the nucleic acid is RNA, the nucleic acid is reverse transcribed to complementary DNA (cDNA) during the amplifying step. In other embodiments, where the nucleic acid is RNA, the nucleic acid is reverse transcribed to cDNA prior to the amplifying step. In some embodiments, the subject is a mammal. In some embodiments, the subject is a human.

[0198]

[0198] Any one of the methods disclosed herein that employ an assay cartridge or detection system for LAMP may be applied to any one of the assay cartridges or detection systems, or both, disclosed herein or those that are previously known in the art. In some embodiments, the assay cartridges or detection systems, or both, comprise elements that permit detection of the electrical signal during the amplifying step. In some embodiments, the electrical signal is impedance or capacitance, or both. In some embodiments, the electrical signal is measured or analyzed compared to a pre-determined control value. In some embodiments, the methods further comprise determining the biological sample as comprising the pathogen, or the gene or genome region thereof.

[0199]

[0199] Any one of the methods disclosed herein may further comprise mixing the biological sample with a reagent and the primer set in the assay cartridge or detection system, or both, prior to the amplifying step. In some embodiments, the reagent is used for LAMP. In some embodiments, the reagent comprises a strand-displacing DNA polymerase and optionally a reverse transcriptase. Some non-limiting examples of strand-displacing DNA polymerases include but are not limited to Klenow fragment, phi29 DNA polymerase, Bsm DNA polymerase, or Bst DNA polymerase, or any combination thereof, or any stranddisplacing DNA polymerase known in the art. Some non-limiting examples of reverse transcriptases include but are not limited to M-MLV reverse transcriptase, reverse transcription xenopolymerase (RTX), or variants thereof, or any combination thereof, or any reverse transcriptase known in the art. In some embodiments, the reagent or the primer set, or both, have been dried and / or lyophilized prior to mixing with the biological sample. In some embodiments, the biological sample is aqueous and dissolves the dried and / or lyophilized reagent or primer set, or both.

[0200]

[0200] Any one of the methods disclosed herein may comprise an assay cartridge or detection system, or both, that comprises a heater. In some embodiments, the amplifying step comprises incubating the biological sample at, optionally a first temperature for a first time period, and at least a second temperature for a second time period. In some embodiments, the amplifying step comprises incubating the biological sample at, optionally a first temperature for a first time period, and one or more second temperatures for one or more second time periods (e.g., a second temperature, a third temperature, a fourth temperature, a fifth temperature, and / or a sixth temperature or more for a second, third, fourth, fifth, sixth, and / or more time periods). In some embodiments, the first temperature is 20°C, 21 °C, 22°C,

[0201] 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C, 30°C, 31 °C, 32°C, 33°C, 34°C, 35°C, 36°C,

[0202] 37°C, 38°C, 39°C, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C, 46°C, 47°C, 48°C, 49°C, 50°C,

[0203] 51°C, 52°C, 53°C, 54°C, or 55°C, or about 20°C, about 21°C, about 22°C, about 23°C, about 24°C, about 25°C, about 26°C, about 27°C, about 28°C, about 29°C, about 30°C, about 31°C, about 32°C, about33°C, about34°C, about35°C, about 36°C, about 37°C, about 38°C, about 39°C, about 40°C, about 41°C, about 42°C, about 43°C, about 44°C, about 45°C, about 46°C, about 47°C, about 48°C, about 49°C, about 50°C, about 51 °C, about 52°C, about 53°C, about 54°C, or about 55 °C, or any temperature within a range defined by any two of the aforementioned temperatures, preferably 23°C or about 23°C or 50°C or about 50°C, and the first time period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 minutes, or about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, or about 15 minutes, or any time period within a range defined by any two of the aforementioned times, preferably 5 to 10 minutes or about 5 to about 10 minutes. In some embodiments, the second temperature or each of the one or more second temperatures (e.g., second, third, fourth, fifth, sixth, and / or more temperatures) is 21 °C, 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C, 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, 41 °C, 42°C, 43°C, 44°C, 45°C, 46°C, 47°C, 48°C, 49°C, 50°C, 51°C, 52°C, 53°C, 54°C, or 55°C, 56°C, 57°C, 58°C, 59°C, 60°C, 61°C, 62°C, 63°C, 64°C, 65°C, 66°C, 67°C, 68°C, 69°C, or 70°C, or about 21 °C, about 22°C, about 23 °C, about 24°C, about 25°C, about 26°C, about 27°C, about 28°C, about 29°C, about 30°C, about 31 °C, about 32°C, about 33°C, about 34°C, about 35°C, about 36°C, about 37°C, about 38°C, about 39°C, about 40°C, about41°C, about 42°C, about 43°C, about 44°C, about 45°C, about 46°C, about 47°C, about 48°C, about 49°C, about 50°C, about 51 °C, about 52°C, about 53°C, about 54°C, or about 55°C, about 56°C, about 57°C, about 58°C, about 59°C, about 60°C, about 61 °C, about 62°C, about 63°C, about 64°C, about 65°C, about 66°C, about 67°C, about 68°C, about 69°C, or about 70°C, or any temperature within a range defined by any two of the aforementioned temperatures, preferably 50°C or about 50°C, and each of the one or more second time periods (e.g., a second temperature, a third temperature, a fourth temperature, a fifth temperature, and / or a six temperature or more for a second, third, fourth, fifth, sixth, and / or more time periods) is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, or 60 minutes, or about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15 minutes, about 16, about 17, about 18, about 19, about 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, or about 60 minutes, or any time period within a range defined by any two of the aforementioned times, preferably 10 minutes or about 10 minutes.

[0204]

[0201] In one embodiment, the first step is performed at or about at 50°C, and the second step is performed at or about at 65 °C. In another embodiment, the first step is performed for about 10 minutes or 10 minutes.

[0205]

[0202] In some embodiments, the amplifying step further comprises incubating the biological sample at a third temperature for a third time period. In some embodiments, the third temperature is 60°C, 61°C, 62°C, 63°C, 64°C, 65°C, 66°C, 67°C, 68°C, 69°C, or 70°C, or about 60°C, about 61 °C, about 62°C, about 63°C, about 64°C, about 65°C, about 66°C, about 67°C, about 68°C, about 69°C, or about 70°C, or any temperature within a range defined by any two of the aforementioned temperatures, preferably 65°C or about 65°C, and the third time period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, or 60 minutes, or about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15 minutes, about 16, about 17, about 18, about 19, about 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, or about 60 minutes, or any time period within a range defined by any two of the aforementioned times, preferably 30 minutes or about 30 minutes. In some embodiments, the first temperature is performed at room temperature (e.g., 23°C or about 23°C) for a time period sufficient to allow the dried down reagents to rehydrate (e.g., 10 minutes or about 10 minutes); the second temperature is performed at 50°C or about 50°C for 10 minutes or about 10 minutes, and the amplification period is then conducted at 65°C or about 65°C.

[0206]

[0203] Enhancement of LAMP can occur when a two-step incubation is used, and that the two-step protocol can result in a lower limit of detection (LOD), than the one-step protocol, with the most consistent detection occurring at the 10 minute time point.

[0207]

[0204] The ramping protocols for LAMP amplification set forth herein (e. g. , the two- step protocol) is suitable for use with any primer directed to any viral or bacterial pathogen nucleic acid template, preferably using the primers described herein, and is not limited to viral avian flu (e.g., H5N1). While it is preferred that the ramping protocols for LAMP amplification set forth herein (e.g., the two-step protocol) are utilized in one or more of the systems described herein (e.g., a be. well® cartridge), it is also evident that the ramping protocols for LAMP amplification set forth herein (e.g., the two-step protocol) will improve the level of detection in other assays that utilize LAMP amplification. Accordingly, more generally, use of a starting temperature that is lower than the amplification temperature of 65 °C for a pre-amplifi cation time period is contemplated to improve the level of detection in LAMP amplification, preferably in one or more of the systems described herein (e.g., a be.well® cartridge) but also in other detection systems that utilize e.g., optical or radioactivity detection. That is, a method of improving a limit of detection of a nucleic acid using LAMP amplification with a primer set at more than one temperature is contemplated.

[0208]

[0205] Any one of the methods disclosed herein may be applied to the detection of a nucleic acid of a pathogen, or a genome region thereof. In some embodiments, the pathogen is a human pathogen. In some embodiments, the primer sets, or the one or more F3 primers, one or more B3 primers, one or more LF primers, one or more LB primers, one or more FIP primers, or one or more BIP primers, or any combination thereof, are designed, configured or selected to be not only specific towards a genome region of a pathogen but also to amplify said specific genome region more efficiently than other primer sets (e.g., more rapidly, exhibiting faster time to detection of a positive amplification and / or with greater specificity). In some embodiments, the pathogen is a virus or a bacterium. In some embodiments, the pathogen is SARS-CoV-2, hepatitis A virus, Influenza A virus subtype H1N1, human immunodeficiency virus- 1, respiratory syncytial virus A, respiratory syncytial virus B, Escherichia coli, Listeria monocytogenes, Mycobacterium tuberculosis, Salmonella enterica, or any combination thereof.

[0209]

[0206] In some embodiments, the primers of any of the methods disclosed herein are provided for LAMP at defined concentrations.

[0210]

[0207] In some embodiments, each of the one or more F3 primers are provided at a concentration of 100 nM, 200 nM, 300 nM, 400 nM, 500 nM, 600 nM, 700 nM, 800 nM, 900 nM, 1000 nM, 1100 nM, 1200 nM, 100 nM, 300 nM, 1500 nM, 1600 nM, 1700 nM, 1800 nM, 1900 nM, or 2000 nM or about 100 nM, about 200 nM, about 300 nM, about 400 nM, about 500 nM, about 600 nM, about 700 nM, about 800 nM, about 900 nM, about 1000 nM, about 1100 nM, about 1200 nM, about 100 nM, about 300 nM, about 1500 nM, about 1600 nM, about 1700 nM, about 1800 nM, about 1900 nM, or about 2000 nM per amplifying reaction, or any concentration within a range defined by any two of the aforementioned concentrations, preferably 200 nM or about 200 nM.

[0211]

[0208] In some embodiments of any of the methods disclosed herein, the method can be multiplexed to be used to detect the presence and / or amount of more than one nucleic acid, such as more than one pathogen, or serotypes, strains, mutants, isolates, species, variants, types, subtypes or clones thereof. In some embodiments, the methods are multiplexed by detecting more than one nucleic acid in a single reaction well 122. In some embodiments, to multiplex the detection, more than one primer sets are included in the single reaction well 122. In some embodiments, the multiplexed methods comprise amplifying the more than one nucleic acid with the more than one primer sets (e.g., in the single reaction well 122) and determining the presence and / or amount of the more than one nucleic acid in the biological sample. In some embodiments, the more than one primer sets comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15,16, 17, 18, 19, or 20 primer sets, each comprising any combination of: one or more FIP primers, and one or more BIP primers. In some embodiments, determining the presence and / or amount of the more than one nucleic acid comprises determining the presence and / or amount of at least one of the more than one nucleic acid in the biological sample without knowing which of the more than one nucleic acid is amplified. In other embodiments, determining the presence and / or amount of the more than one nucleic acid comprises determining the presence and / or amount of each of the more than one nucleic acid in the biological sample.

[0212]

[0209] In some examples, LAMP primers specific for avian influenza can be suitably used.

[0213] Tracking Infection Potential

[0214]

[0210] Various conditions exist under which knowledge regarding whether a subject or product is infected, is at risk of being infected, or was exposed to an individual or other products that have been infected are useful. For example, during a local epidemic or global pandemic, a store or public venue that provides services, entertainment, or goods to the public may limit entry to people that are determined to be infected or have a likelihood of being infected or are at risk of being infected (e.g., people having a wellness score that deviates from a threshold value commensurate with that of healthy individuals) in favor of people that are not infected, have a reduced likelihood of being infected, or are not at risk of being infected (e.g., people having a wellness score commensurate with a range for healthy individuals). The store or public venue may implement restrictions to entry for individuals that have wellness scores, which deviate from the healthy threshold value (e.g., outside of the range for healthy individuals) so as to reduce the risk of exposing other members of the population to a pathogen. In particular, during cold and flu (influenza) season or localized outbreaks of contagious pathogens (for example, the COVID-19 coronavirus, tuberculosis, and the like), various entities (for example, people, stores, restaurants, business, government agencies, and so forth) may wish to limit access to spaces, facilities, services, and so forth to people who are not or likely not infected. Therefore, tracking wellness scores that can provide information specific and relevant to a particular infection of interest for large numbers of people in a population is desired.

[0215]

[0211] Similarly, with respect to infected animals or products in the food supply, e.g. , cattle, chickens, turkeys, lamb, fish, vegetables, fruits, or juices, or milk products, it is beneficial to evaluate the health or contamination status of the animal or product, determine whether it is infected with a pathogen, and / or determine whether the animal or product poses a risk to other animals or products in the population or to consumers. By evaluating the wellness scores for animals in an animal population or products in the food supply and comparing these wellness scores to thresholds indicative of healthy or diseased individuals or contaminated or non-contaminated products, one can choose to isolate or remove animals or products having wellness scores indicative of infection by a pathogen from animals having healthy wellness scores in the population or uncontaminated products in the food supply so as to reduce the spread of disease within the animal population, cross-infection to humans, or contamination of the food supply.

[0216]

[0212] Initially identifying the healthy or uninfected subj ects from those subj ects that are infected is difficult, especially when, generally, the identification of infected subjects requires specific testing procedures and diagnostic tests. Such limitations would often hamper testing and monitoring capabilities in previous systems. For example, taking a test for a particular infection and obtaining the results for the test can involve going to a clinic or medical professional’s office, waiting for the test to be administered and processed, and waiting for the results to be available and reported to the individual. In many instances, such testing processes take many days to complete. Even then, various risks with the tests exist, such as cross contamination of the test samples, mix-up of tests or test results, and so forth. Accordingly, such testing may not be practical or even possible for use in determining whether to permit or deny entry of people into a location, event, and so forth.

[0217]

[0213] Furthermore, in some instances, testing alone may not be indicative of infection or infectivity to others, because other factors may contribute to a likelihood of a subject or product being infected or contaminated. Examples of these factors include the subject or product traveling to locations known to be hot spots of infection, not taking proper precautions when in public or in product handling when risk of infection is high, having been in contact with someone confirmed to be infected, having shown symptoms consistent with being infected, and so forth. Such factors, in conjunction with test results and corresponding information, may be applicable or useful for determining whether to allow entry or access of a product, person or animal to a location or into the food supply.

[0218]

[0214] Additionally, contact tracing for people, products, or animals determined to be infected or possibly exposed to infected subjects or products is reliant on various factors and can be difficult to perform with high confidence and accuracy. For example, previous general contact tracing methods for people, products, or animals can include identifying a subject or product that is infected or contaminated, contacting that subject (or a responsible subject) or evaluating the chain of custody of a product to determine whether the subject or product may have exposed others or where the subject or product was exposed, attempting to identify other subjects or products that were at the same location or in the vicinity of the infected subject or product during the infectious period, and then attempting to identify and / or contact other potentially infected subjects or products. Thus, conventional general contact tracing methods often rely on a subject’s self-reporting, memories, and willingness to participate, record keeping capabilities, and / or communication capabilities, which often can be difficult to implement.

[0219]

[0215] The systems and methods described herein improve both determining whether a subject or product is infected and contact tracing. More specifically, the systems and methods described herein provide assistance in determining whether to allow entry or access to locations, the food supply, or the general population of individuals based on generally accepted indicators of infection while maintaining appropriate and consistent application of the indicators and analysis, maintaining privacy and confidentiality, and providing ongoing data collection and updates. For example, the systems and methods described herein may provide and utilize profiles, markers, identifiers, or accounts associated with people, animals, or products to establish a wellness score for the person, animal, or product based on which people, animals, or products are allowed or deny entry or access to locations, populations of other animals, or the food supply. A person’s, animal’s, or product’s identity may be confirmed using, e.g., biometrics, bar coding, QR coding, RFID coding with or without other authenti cation systems to ensure the subject or product being presented for entry or access to a public place, general population, or food supply is confirmed to be the subject or product for which the wellness score applies.

[0220]

[0216] For example, the individual or product may create a profile or have a profile created specific to the subject or product. In some embodiments, the subject’s profile may be integrated with various medical accounts or health status records for the subject such that the subject’s medical history is accessible via the profile and can be accounted for in any corresponding analysis. By linking or integrating the subject’s medical accounts or health status records, any test results, health metrics, identification information, authentication information (for example, biometrics information and the like), appointments, and / or other corresponding information from the medical accounts can be considered when generating the wellness score for the subject. The subject’s profile may include various fields, including identifier information (which may comprise a unique identifier), user credentials (for example, user name and passwords or similar identifying information), health information, test results, locations visited, health surveys, contact tracing information, the wellness score, and so forth.

[0221]

[0217] The system may generate the wellness score based on one or more algorithms that dynamically account for the information in the subject’s or product’s profile. For example, age of test results (for example, time since the last test taken) has a varying weight in the algorithm, such that as the test results age (i.e., as the test results become older or are further in the past), the test results may lose weight in the algorithm. This allows more recent activity in the profile to have a greater impact on the subject’s or product’s wellness score. Furthermore, the test results may have a higher weight than other fields in the profile. In some embodiments, the algorithm may account for relationships between users or products when generating wellness scores. For example, if a first user or product is part of a family unit or unit of products and comes in contact with both a second user or product and a third user or product, then the algorithm for generating the wellness score accounts for such relationships or contacts. Thus, if the first and second users or products of the family unit or product unit have taken tests or have been tested with results that show that the first and second users or products are not infected, then the algorithm may generate the wellness score for the third user or product accounting for the recent, not infected test results for the first and second users or products even if the third user or product was not recently tested. This may be because the living situation or contact situation and the particular infection of interest indicates a high likelihood that if a majority of subjects in a household or animal population or products are not infected, then it is likely that none of the subjects or products are infected, and so forth. The algorithm(s) may generate the wellness score to indicate how the various fields are related to an expected wellness (or likelihood of being infected) for the subject or product. In some embodiments, the wellness score is indicative of an expected likelihood that the subject or product is infected. For example, the wellness score is inversely related to the expected likelihood that the subject or product is infected, where a high wellness score is indicative of a low likelihood of being infected and a low wellness score is indicative of a higher likelihood of being infected.

[0222]

[0218] The system may interface with various external systems or devices, for example, to enable one to access their information and control access to information in the profile. The system may also interface with testing devices (for example comprising one or more of the readers and cartridges described herein), medical devices, computing systems, or third-party devices. The third-party devices may comprise computing systems or devices used by third party entities that provide goods or services or at which multiple people may gather, such as stores, restaurants, event venues, and so forth. The third-party entities may use the third-party devices (for example, site devices) to identify whether the subject or product has a wellness score that exceeds a threshold value (e.g., has a likelihood of being infected that falls below a threshold). For example, when the subject or product arrives at the third-party location, the subject may present identification or the product can be identified by an identifier, marker, code, and / or biometric information, via the third-party device, to authenticate the subject or product. The third-party device may convey the identifying or biometric information with the subject’s or product’s identifier to the system, which uses the identifying or biometric information to authenticate the subject’s or product’s identity. If the identity is authenticated or verified, then the system returns, to the third-party device, the subject’s or product’s wellness score from the subject’s or product’s profile. In some embodiments, the third-party system may use the subject’s or product’s wellness score for comparison against the third party’s threshold requirements. Alternatively, the system may compare the subject’s or product’s wellness score against the third party’s threshold requirements or other standard threshold requirements and return an indication regarding whether or not the subject or product is allowed to access the third-party location or not. As such, third parties are able to determine whether people, animals, or products can access locations accurately based on unbiased, scientific information for a specific individual or product. Alternatively, or additionally, the system can provide the individual or product with a computer-generated code that is indicative or representative of the individual having the wellness score that meets or exceeds the threshold for entry to the third-party location. For example, the system can generate a barcode, radio-frequency identification (RFID) code, or quick response (QR) code that the individual or product can present at the third-party location and which the third party can scan to determine that the individual or product can enter the location.

[0223]

[0219] In some embodiments, the system and methods update the subject’s or product’s profile dynamically with additional information, for example when the subject or product takes a new test, is tested, or visits a health professional. Thus, when the subject or product takes the new test, or is newly tested the system updates the subject’s or product’s profile based on results from the new test and then uses these results to revise the wellness score. Thus, as information in the profile changes, the system may update the subject’s or product’s wellness score accordingly such that the subject’s or product’s wellness score is always up-to-date.

[0224]

[0220] Accordingly, the systems and methods described herein, incorporated with the cartridges and reader devices of the platform also described herein, provide for consistent and accurate determinations of whether people, animals, or products are infected while maintaining privacy for the people and granular monitoring of animals and products especially in the food supply. Further details are provided herein.

[0225]

[0221] An example of such a system is shown in FIG. 8, which shows a network diagram of a system 800 for tracking users’ wellness scores and likelihood of being infected. In some embodiments, the system 800 operates over a network 805 and comprises a user device 810, a testing device 815, a site device 820, a profile database 825, and a server 830. The system 800 may implement the methods described herein.

[0226]

[0222] The network 805 may allow various computing devices and / or other electronic devices to communicate with each other via wired or wireless communication links. The communications links between the various computing devices and components of the system 800 may be wired or wireless connections (or a combination thereof) and may be part of a secured or unsecured network, such as a local area network (LAN), a wide area network (WAN), a combination of networks, and / or the Internet. For example, the communication links may be achieved through one or more a Wi-Fi system, Bluetooth® wireless technology, Ethernet, cellular communications, or satellite communications, or the like.

[0227]

[0223] In some embodiments, the user device 810 comprises an external computing device. The user device 810 comprises a user interface through which the user may interact with various components in the system 800, for example the testing device 815, the site device 820, the profile database 825, and the server 830. The user device 810 may comprise any computing device used by the user. For example, the user device 810 comprises one or more of a smart phone, a tablet, a smartwatch, a computer, a laptop, or any similar mobile or stationary computing device. In some embodiments, the user device 810 communicates directly with the testing device 815 (e.g., without communicating through the network 805), for example via one of the wired or wireless communication links. The user device 810, as described above, may allow the user to manage the testing device 815 (comprising taking and reviewing results from the test) and respond to health information surveys (for example, provide information or responses, temperature measurements, and the like). The user of the user device 810 may also use the user device 810 to review and manage the user’s profile, an animal’s profile, or a product’s profile as stored in the profile database 825. Additionally, the user device 810 enables the user to obtain and monitor the wellness score in the user’s profile an animal’s profile, or a product’s profile. The user may use the wellness score to generate a barcode, QR code, RFID code, or other computer-generated value to present to third parties to identify whether the user, animal, or product is infected. In some embodiments, the user may use the wellness score to combine with other information, for example a ticket barcode or number for a ticket for admission of entry to an event or location (for example, a plane or transportation ticket, event ticket, and so forth). The user can then present the combined barcode or number (for example, via the user device 810) to the site device 820 to indicate both payment of appropriate fees as well as the user’s wellness score indicative of the user being infected or not. Thus, the single barcode or computer readable number may be used with the third-party site device 820 to gain admission to a location or event. Further details are provided below with respect to the description of the site device 820 and FIG. 9. In some embodiments, the user device 810 operates an application (or similar software code) associated with the system 800. The user device 810 may use the application to track information, for example location information of the user device 810 and identification information for other user devices 810 with which the user device 810 comes into close proximity. The user device 810 may store the location information and identification information in the user profile for the user in the profile database 825. In some embodiments, the user profile also enables the individual to monitor health information and records provided by various health institutions, for example the individual’s doctor, records from blood tests, and the like, for example via the user device 810.

[0228]

[0224] In some embodiments, the user device 810 may locally store test results from an infection test and enable the user to present the test results to the site device 820 to gain entry to an event or to pass the animal or product on to a subsequent location. For example, the user takes the test using the testing device 815 for a pathogen of concern and the test results are stored on the user device 810 (and optionally stored in the user profile in the profile database 825). The user may then take the user device 810 to the event location and present the user device 810 to the site device 820. Alternatively, test results for a tested animal or product can register on a user’s device, which can be presented at a subsequent location during transport of the animal or product. The site device 810 may access the test results via the computer-generated code or directly via a pass / fail (not infected / infected) indication on the user device 810. In some embodiments, the user may provide an identifier and / or biometrics information (via one of the site device 820 or the user device 810) to authenticate that the test results provided on the user device 810 actually belong to the user presenting the user device 810 or the animal or product for which the test results apply. For example, logging into an application on the user device 810 that shows the test results may require a biometric input as opposed to just the username and password. As described herein, user credentials may include or comprise the username and password information and / or biometrics information. As such, the user device 810 may comprise the application that, when used in conjunction with a user profile or testing device 815, requires biometric inputs from users and associates test results and profile information with the biometric inputs and / or the user identifier information or identifying information about the tested animal or product so that test results cannot be shared with third parties or between users or event or location moderators and are always associated with a single user, animal or product.

[0229]

[0225] The testing device 815 may comprise a combination of a cartridge (for example, one of the test cartridges described herein, for example the test cartridge 100, and so forth) and a reader device (for example, the reader devices or analyzer 300), as introduced above. The testing device 815 may test for or identify pathogens, genomic materials, proteins, and / or other small molecules or biomarkers infections, diseases, and so forth, which may be indicative of whether or not the individual or product is infected, based on the test cartridges and readers described herein. As described above, the testing device 815 communicates with the user devices 810. However, the testing device 815 may optionally communicate directly with the network 805 and, therefore, communicate with one or more of the profile database 825 and the system server 830 via the network 805 or via the user device 810 in addition to the user device 810. The testing device 815 may enable testing of biological samples from the individual or product and may provide the results from the testing to the individual (via the user device 810), the profile database 825, or a third party. Additionally, the testing device 815 may perform diagnostic tests, including tests that utilize test strips, diagnostics screenings, tests that require biological samples, and so forth. Further details regarding the testing device (more specifically, the reading device and the cartridge) are provided throughout this disclosure.

[0230]

[0226] In some embodiments, the user may obtain or take one or more tests via the testing device 820 to determine if the user, animal, or product is infected, for example with a viral or bacterial infection. For example, the user may employ an assay or similar cartridge and the reader device of the testing device 820 to perform the test. The testing device 820 may be used to detect whether the user, animal, or product is infected by detecting specific genomic material from a sample from the user, animal or product that the user deposits into the cartridge of the testing device 820. The user then inserts the cartridge into the reader device, which analyzes the sample to detect whether the sample indicates that the user, animal, or product is infected. Further details regarding how the cartridge and reader device operate to detect whether the sample indicates that the individual, animal or product is infected are provided herein. In some embodiments, the user or animal is infected whenever the sample from the user includes a marker associated with the infection or corresponding pathogen, regardless of whether the user is asymptomatic or symptomatic.

[0231]

[0227] The site device 820 may comprise a computing system or similar device that accesses the system 800 via the network 805 to enable access to information from the system 800 at a location separate from the testing device 815 and / or the user device 810. The site device 820 may be operated by the third party for use to monitor and / or manage entry of people, animals or products to a location or event or into the food supply. The site device 820 includes a user interface to allow the users to provide identifiers and credentials (for example, username and password or biometric information) and view results of comparing the wellness score with the threshold. For example, the location can be an airport, a restaurant, an event venue (e.g., a concert, party or sports event), or in the case of animals a storage or processing facility, or in the case of products, a packing facility. In some embodiments, the site device 820 comprises a barcode reader, QR reader, RFID reader, or similar computer-generated code reader to read the computer-generated code described herein.

[0232]

[0228] In some embodiments, the site device 820 may operate in conjunction with the testing device 815 to allow tests to be administered at the location, e.g., an event, and then compared or aggregated with information from the user profiles stored in the profile database 825. For example, the user attending the event at the location or an animal or product arriving at a location are tested using the testing device 825 at the location and the site device 820 may obtain other user profile information to generate the wellness score, also including the test taken at the location using the testing device 825, to generate an updated wellness score while the user, animal or product is at or in a vicinity of the location (e.g., at a pre-admittance point). The site device 820 may then compare the wellness score with the threshold score for entry to determine whether to allow or deny entry of the user, animal, or product. As such, entry into the location for the user, animal or product may be based on the test that is as recent as possible relative to the event, location, or transport of the animal or product, as well as, the profile information for the user, animal or product where biometrics information or other identifying information is used to access the profile information from the profile database 825 when the biometrics information or other identifying information matches that of the user’s profile (identified by the identifier) or the profile for the animal or product being tested. The site device 820 may then store the updated wellness score in the user profile, or profile for the animal or product in the profile database 825.

[0233]

[0229] The server 830 may allow the site device 820 to access the wellness score for the user, animal or product that presents to gain entry to the location or event. For example, the third party, when using the site device 820 to manage entry of people, animals or products into the location or event, applies or establishes the threshold wellness score that people, animal or products must meet or exceed to be granted entry. In some embodiments, the threshold score is set by the third party or by a larger body (for example, governing body). The site device 820 may be used to identify the wellness score for users, animals or products that are presented for entry and compare the wellness score to the threshold score. For example, the site device 820 obtains the wellness score for each user, animal, or product in various ways. For example, the site device 820 may obtain identifying and / or biometric information from the user, animal or product and submit the obtained information to the server 830 for authentication of the user, animal or product via the user’s, animal’s or product’s profile in the profile database 825. Thus, the site device 820 may be configured to capture and convey identifying information and / or biometric information for the user, animal or product for example user fingerprints, photo or video based biometrics (for example, facial recognition or retinal scans), user physiological based biometrics (for example, facial recognition, hand geometry recognition, finger geometry, iris / retinal scanning, palm vein recognition, ear recognition, and so forth), user voice recognition (for example, cadence, voice patterns, and so forth), user writing, signature, or typing pattern recognition, biological sample recognition, movement or gait recognition, bar coding, QR coding, RFID coding, branding, and the like. The server 830 may confirm whether the identifying and / or biometric information match a user, animal or product profile in the profile database 825. If they match, the server 830 may identify the wellness score in the user, animal or product profile and return it to the site device 820. If they do not match, then the server 830 may return an error or request replacement identifying and / or biometric information. When the site device 820 receives the wellness score for the user, animal or product, the site device compares the received wellness score with the threshold score and determines whether the user, animal or product can enter a location (for example, when the wellness score exceeds the threshold, the user, animal or product can enter and when the wellness score does not exceed the threshold, the user, animal or product cannot enter). Alternatively, the site device 820 may scan a computer-generated code for each user, animal or product seeking entry and parse the scanned code to identify the wellness score for the user, animal or product. The site device 820 may then compare to parsed wellness score to the threshold score, as described above, to determine whether the user, animal or product is allowed entry. In some embodiments, the site device 820 further provides details of what users, animals or products (for example, via user, animal or product identifiers) were granted entry by the site device 820 to the server 830. This information may be used by the server 830 for contact tracing purposes so that the server 830 may identify all users, animals or products at the location at any given time. Thus, the site device 820 may determine to limit entry to the location and obtain contact tracing information.

[0234]

[0230] The profile database 825 may store profile information received over the network 805 and provide responses to requests for information received via the network 805. For example, as described above, the profiles in the profile database 825 may comprise, for each user, animal or product having a profile stored therein, one or more fields, including identifier information (for example, user name and passwords or similar identifying information), health information, testing information and results (for example, including when and what tests were taken, the corresponding results, and / or titers available), locations visited, health surveys, contact tracing information, the wellness score, and so forth. The profile database 825 may update profiles for users, animals or products as new test results, health information, or corresponding information is provided to the profile database 825 with respect to corresponding user identifiers. Additionally, the profile database 825 may store updated wellness scores that are generated by the server 830, as described herein. As described above, the profile database 825 may provide the wellness score for the user, animal or product based on receipt of a request for the wellness score when the request includes matching user identification information and biometric information. The biometric or identifying information stored in the user, animal or product profile may include user fingerprints, photo or video based biometrics (for example, facial recognition or retinal scans), physiological based biometrics (for example, facial recognition, hand geometry recognition, iris / retinal scanning, palm vein recognition, ear recognition, and so forth), voice recognition (for example, cadence, voice patterns, and so forth), writing, signature, behavioral recognition, typing pattern recognition, physical movement recognition, navigation patterns, biological sample recognition, QR coding, RFID coding, bar coding, branding and the like. This biometric and / or identifying information may be updated in the profile database 825 based on updated information received from the user, animal or product and so forth.

[0235]

[0231] For example, the server 830 may manage requests for wellness scores and other user, animal or product profile information received from the site device 820. Additionally, the server 830 may determine the wellness scores for user, animal or product profiles based on the other information stored in the user, animal or product profile according to one or more algorithms. For example, the one or more algorithms may incorporate freshness of medical information into the wellness scores by reducing values for aged information (for example, reducing the wellness score generated based on information in the user’s profile even when there is no change to the information except with respect to its freshness. This may be because as the information in the user’s, animal’s, or product’s profile ages, the corresponding likelihood that the profile information is missing relevant or potential infection indicating data increases. Thus, a first user, animal, or product profile with information that has aged a month (i.e., has not been updated in a month) may have a substantially lower wellness score as compared with a second user, animal, or product profile having information updated that has aged two days (i.e., has not been updated in two days), even if the information between the user’s, animal’s, or product’s profile is identical with exception to the age of the information. In some embodiments, the server 830 enables operational use of the system 800, for example controlling authentication by the site device 820 and user device 810 of user, animal, or product profile identifying and / or biometrics information or user access or storage of data in their profiles in the profile database 825.

[0236]

[0232] For example, in some embodiments, the server 830 may use the location information and identification information collected by the user device 810 to perform contact tracing as appropriate and needed. For example, the server 830 may receive an indication that a first user, animal, or product is infected with a pathogen (for example, from testing with the testing device 815 or information added to the user’s health information records). The server 830 may then use the contact tracing information for the first user’s user device 810 or the animal or product profile to identify a second user, animal or product (and any additional users, animals or products) that came into close proximity with the first user, animal or product for example based on the stored location information and identifying information. Based on this information, the server 830 may contact the second user or managers for the animals or products (and any additional users or managers for the animals or products) regarding the possible exposure and suggest that appropriate steps, such as take a corresponding test using the testing device 815, visit the medical professional, quarantine, or removal form the transportation location and so forth.

[0237]

[0233] In some embodiments, one or both of the server 830 and the user device 810 interface with the user’s calendar or events to which the user is expecting to go (for example, a scheduled concern, flight, and so forth). In view of the expected event and need to meet the threshold wellness score to be assured entry, the server 830 and / or the user device 810 may provide instructions to the user regarding when to take the test, when or where to avoid traveling or visiting in advance of the event, and so forth. The instructions may be generated, based at least in part, on the algorithm(s) used to generate the wellness score in the user profiles. Thus, based on the algorithm(s), the server 830 or the user device 810 may determine which factors of the user profile the user can impact through action (for example, take a new test, avoiding stores between certain hours, and so forth). For example, the server 830 and / or the user device 810 instructs the user to take a test at least one day before the event to ensure the user’s profile and wellness score are appropriately updated with the latest information but no longer than seven days before the event to avoid aging of the profile information. Thus, the algorithms described herein and as applied by the systems and methods described herein, may proactively assist users to help ensure they gain entry to the scheduled event.

[0238]

[0234] In some embodiments, the server 830 may integrate with external profiles, for example travel profiles, credit card profiles, payment services profiles, and the like. Such integration may allow the wellness score to be integrated with event information or the like. For example, if the individual purchases a plane ticket or event ticket, the server 830 of the system 800 may integrate the wellness score QR code described above (or corresponding information) with a QR code for the individual’s plane or event ticket QR code or other barcode or identifier into a combined identifier. The combined identifier may represent the individual’s wellness score, as well as, confirmation of payment for the flight or event. In some embodiments, instead of integrating with the external profiles, the server 830 may provide sufficient information to enable the individual to manually integrate the wellness score QR code information with ticket barcode information (for example, via the ticket vendor, and so forth).

[0239]

[0235] In some embodiments, the system 800 may use biological samples from a surface (for example, a high contact surface), an environment (for example, a high-risk environment such as an infection testing location or medical professional’s office), or an object to determine whether the surface, environment, or object was contaminated by a pathogen. For example, the third-party entity may use the testing device 815 to determine whether the location may have been contaminated during the event and communicate that information to the server 830 so that any users that visited the location during the event can be notified, via their user profiles and stored location information, of a possible exposure by visiting the location.

[0240]

[0236] As described above, the networked system 800 may enable the communications and interactions between the various components of the system 800, for example the user device 810, the testing device 815, the site device 820, the profile database 825, and the server 830. FIG. 9 is a flow diagram 900 showing example interactions between components of the networked system 800 of FIG. 8. The communications between these components may occur via one of the communication links described herein.

[0241]

[0237] Again, with reference to FIG. 9, the profile database 825 may create the user profile for the user using system 800. Additionally, the interactions shown in FIG. 9 assume that the user has logged or signed into the user profile and / or the corresponding application on the user device 810 that allows the user to take the test with the testing device 815. The testing device 815 may receive a user input at 901, the user input comprising the user’s biological (or other) sample to deposit in the testing device 815 (for example, via the cartridge). The testing device 815 may perform the corresponding test of the provided sample and generate test results, as described herein using the cartridge and the reader of the testing device 815. These test results may then be communicated to the user device 810, at 910, and / or conveyed directly to the profile database 825 via the network 805. In some embodiments, before receiving the test results at 905, the user provides biometric information and / or other identifying information or credentials to indicate an association with the test results and to indicate that the test results are to be associated with the user (and the user’s profile).

[0242]

[0238] The user device 810 may further receive a user input 910 that comprises one or more responses to health inquiries or surveys, temperature measurements, and so forth, that corresponds to health information for the user. The received test results from 905 and the received responses or other health information from the user input 910 are stored in the profile database 825 at 915. In some embodiments, the test results from 905 and the health information from the user input 915 are stored locally at the user device 810 to enable the user device to process information and generate the wellness score.

[0243]

[0239] The profile database 825 may receive the information from the user device 810 and / or the testing device 815 and store the information in the profile database 825 in the user profile associated with the user. Where the user profile already exists in the profile database 825, as described above, the profile database 825 may update the user profile and calculate the wellness score at 920. Where the user profile does not already exist in the profile database 825, the profile database 825 may create the user profile, store the health information from the user device 820 in the profile database 825, and store the test results from the testing device 815 in the profile database 825, in the created user profile. Additionally, though not shown in FIG. 9, the user may provide credentials or authentication information to authenticate that the user is affiliated with or associated with the user profile. For example, the interaction between the user device 810 and the profile database 825 may include the user’s identifying information or credentials or authentication information (for example biometrics information or username / password information). The profile database 825 may associate each user profile with a particular user via the identifier and the biometrics information or username / password information. Thus, when the user provides the identifier and the biometrics information or credentials, the profile database 825 may know which user profile to update based on the identifier and confirmation that the biometrics information or credentials are associated with the received test results and health information from 915. Thus, at 920, the profile database 825 may determine which user profile to update based on the biometrics, identifying information, or credentials received from the user and update the corresponding user profile accordingly.

[0240] As described herein, the user that provides the user input 901 and the user input 910 may travel to a location using the site device 820 and provide a user input 925 to the site device 820. The user input 925 may comprise the identifier and credentials or biometric information to show that the user is the subject that the user claims to be. The credentials help ensure that the user’s confidentiality is maintained and that all information associated with the user is properly affiliated with the user’s profile. The user input 925 is communicated to the server 830, such that the user’s identifier, credentials, and / or biometrics information are conveyed to the server 830 along with a request for the user’s wellness score. The site device 820 may request the user’s wellness score to determine whether the user can be allowed to enter the site or location of the site device 820. In some embodiments, though not shown in FIG. 9, the site device 820 may communicate directly with the profile database 825. Instead, as shown in FIG. 9, the server 830 acts as an intermediary between the site device 820 and the profile database 825. In some embodiments, the server 830 operates to authenticate the site device 820 to determine that the site device 820 is allowed to access information from the profile database 825 and use the system 800.

[0244]

[0241] When the server 830 confirms that the site device 820 is authenticated to use the system 800 and access information from the profile database 825, the server 830 passes the user credentials and score request to the profile database 825 at 935. Based on the received user credentials and score request, the profile database 825 may use the information in the user credentials and score request to confirm that the user is affiliated to the user profile identified by the user credentials at 940. When the user credentials do match the user for the user profile, then the profile database 825 may identify the wellness score and return the wellness score to the server 830, which the server 830 returns to the site device 820 at 950. When the profile database 825 communicates with the site device 820 directly, then the profile database 825 returns the wellness score directly to the site device 820, though not shown in FIG. 9.

[0245]

[0242] The site device 820 may compare the received wellness score to a threshold score value at 955. This comparison, as described herein, may be used to determine whether the user is allowed or denied entry to the location or event. The site device 820 then allows or denies entry for the user to the location at 960. In some embodiments, the comparison of the wellness score and the threshold may occur at the server 830.

[0243] FIG. 10 depicts a general architecture of a computing device implementing one or more of the components of the system of FIG. 8. The general architecture of the computing system 1000 depicted in FIG. 10 includes an arrangement of computer hardware and software that may be used to implement aspects of the present disclosure. The hardware may be implemented on physical electronic devices, as discussed in greater detail below. The software may be implemented by the hardware described herein. The computing system 1000 may include many more (or fewer) elements than those shown in FIG. 10. It is not necessary, however, that all of these generally conventional elements be shown in order to provide an enabling disclosure. Additionally, the general architecture illustrated in FIG. 10 may be used to implement one or more of the other components illustrated in FIG. 8.

[0246]

[0244] As illustrated, the computing system 1000 includes a processing unit 1090, a network interface 1092, a computer readable medium drive 1094, and an input / output device interface 1096, all of which may communicate with one another by way of a communication bus 1070. The network interface 1092 may provide connectivity to one or more networks (for example, the network 805) or computing systems (for example, the any of the components of the system 800). The processing unit 1090 may thus receive information and instructions from other computing systems or services via network 805. The processing unit 1090 may also communicate to and from primary memory 1080 and / or secondary memory 1098 and further provide output information for an optional display (not shown) via the input / output device interface 1096. The input / output device interface 1096 may also accept input from an optional input device (not shown).

[0247]

[0245] The primary memory 1080 and / or secondary memory 1098 may contain computer program instructions (grouped as units in some embodiments) that the processing unit 1090 executes in order to implement one or more aspects of the present disclosure. These program instructions are shown in FIG. 10 as included within the primary memory 1080 but may additionally or alternatively be stored within secondary memory 1098. The primary memory 1080 and secondary memory 1098 correspond to one or more tiers of memory devices, including (but not limited to) RAM, 3D XPOINT memory, flash memory, magnetic storage, cloud storage objects or services, block and file services, and the like. In some embodiments, all of the primary memory 1080 or the secondary memory 1098 may utilize one of the tiers of memory devices identified above. The primary memory 1080 is assumed for the purposes of description to represent a main working memory of the computing system 1000, with a higher speed but lower total capacity than secondary memory 1098.

[0248]

[0246] The primary memory 1080 may store an operating system 1084 that provides computer program instructions for use by the processing unit 1090 in the general administration and operation of the computing system 1000. The memory 1080 may further include computer program instructions and other information for implementing aspects of the present disclosure. For example, in one embodiment, the memory 1080 includes a user interface unit 1082 that generates user interfaces (and / or instructions therefor) for display upon a computing device, e.g., via a navigation and / or browsing interface such as a web browser or software application installed on the computing device.

[0249]

[0247] The computing system 1000 of FIG. 10 is one illustrative configuration of such a device, of which others are possible. For example, while shown as a single device, the computing system 1000 may, in some embodiments, be implemented as multiple physical host devices. In other embodiments, the computing system 1000 may be implemented as one or more virtual devices executing on a physical computing device. While described in FIG. 10 as a computing system 1000, similar components may be utilized in some embodiments to implement other devices shown in the system 800 of FIG. 8.

[0250] Dry Reagents

[0251]

[0248] Traditionally, nucleic acid amplification and detection techniques, as described herein, involve various steps, including multiple pipetting steps for preparing for the amplification and detection techniques. Additionally, these techniques also often involve setting up various reagents (or reagent mixes), such as enzymes, primers and dNTP, for use in the amplification and detection techniques. Any of these steps can introduce errors, which can, in turn, lead to error-prone results in the amplification and detection. Additionally, the reagents may need a cold-chain compartment or facility to maintain stability of the reagents during storage and transportation. However, as described above, the systems and methods described herein utilize a dried, pre-optimized enzymatic reagent or reagent mix in the test cartridge, which overcomes these issues. The dried reagent, which is stable and ready-to- use for nucleic acid amplification, can be safely stored and transported in or via the test cartridges for use on demand with one of the readers described herein. As also described herein, when used to detect pathogens, genomic materials, proteins, and / or other small molecules or biomarkers, the test cartridge receives a sample deposited into a well (for example, via a swab or pipette). The sample is then mixed with the dried reagent(s) in the cartridge when the cartridge is inserted into the reader, thereby rehydrating the dried reagent(s) to further the testing. Three different improvements to the testing process are provided below to improve the general amplification and detection techniques. In accordance with the present disclosure, BSA can be included in the dry reagents. Drying down BSA may be desirable, as dry BSA is relatively stable over long periods of time. In drying down the reagents, BSA can be included at 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.10%, 0.11%, 0.12%, 0.13%, 0.14%, 0.15%, 0.16%, 0.17%,

[0252] 0.18%, 0.19%, 0.20%, 0.21%, 0.22%, 0.23%, 0.24%, 0.25%, 0.26%, 0.27%, 0.28%, 0.29%,

[0253] 0.30%, 0.31%, 0.32%, 0.33%, 0.34%, 0.35%, 0.36%, 0.37%, 0.38%, 0.39%, 0.40%, 0.41%,

[0254] 0.42%, 0.43%, 0.44%, 0.45%, 0.46%, 0.47%, 0.48%, 0.49%, 0.50%, 0.51%, 0.52%, 0.53%,

[0255] 0.54%, 0.55%, 0.56%, 0.57%, 0.58%, 0.59%, 0.60%, 0.61%, 0.62%, 0.63%, 0.64%, 0.65%,

[0256] 0.66%, 0.67%, 0.68%, 0.69%, 0.70%, 0.71%, 0.72%, 0.73%, 0.74%, 0.75%, 0.76%, 0.77%,

[0257] 0.78%, 0.79%, 0.80%, 0.81%, 0.82%, 0.83%, 0.84%, 0.85%, 0.86%, 0.87%, 0.88%, 0.89%,

[0258] 0.90%, 0.91%, 0.92%, 0.93%, 0.94%, 0.95%, 0.96%, 0.97%, 0.98%, 0.99%, or 1% or within a range defined by any two of the previous values, though in some cases values outside of these ranges may be suitable.

[0259] Mixing

[0260]

[0249] In the process described above, rapid rehydration of the reagent(s) by the sample and effective mixing with the sample is necessary for good assay performance (for example, accurate and consistent test results), which may include high sensitivity and amplification efficiency. While passive diffusion or mixing of the reagent with the sample and heat energy for rehydration may result in sensitivity and amplification efficiency that meets a minimum threshold, such a mixing strategy may, in some instances, not be sufficiently effective. Improved methods or strategies may include introducing various components to improve mixing, for example mixing beads, which may be magnetic.

[0261]

[0250] In some alternatives, magnetic beads (or similar magnetic or mixing objects of non-magnetic materials, hereafter magnetic beads) inserted into a mixing chamber (for example, the mixing well 224 or test well 1258) may enhance the rehydration and mixing of the reagents with the sample. The magnetic beads may exist in the mixing chamber of the test cartridge when packaged with the reagents (for example, the magnetic beads may be inserted into the mixing chamber when or prior to when the reagents are inserted or injected into the mixing chamber). In some instances, the magnetic beads are movable by shaking the test cartridge (for example, the user shaking the reader once the test cartridge with the sample is inserted into the reader). Alternatively, or additionally, the user can move the magnetic beads by a magnetic field, sonic waves, oscillations, vibrations, sonication, shaking, or similar contact or non-contact methods to ensure the improvement to the rehydration and mixing between the sample introduced into the test cartridge and mixed with the reagent. In some embodiments, these fields, forces, and movements are generated by one or more of a magnetic field generator (for example a permanent magnet or an electromagnet), a vibration generator, a sonic generator, and physical movement In some embodiments, the reagent may be a dry or dried reagent or a liquid reagent.

[0262]

[0251] The movement of the magnetic beads will help mix and combine the reagent with the sample, similar to physical mixing devices (mixers, stirring devices, and so forth) assist in mixing larger quantities of mixture. The analyzer may employ one or more corresponding components that cause the magnetic field (for example, an electromagnet) or other fields by which the magnetic beads are moved within the mixing chamber of the test cartridge. Thus, the analyzer may cause the magnetic beads in the test cartridge to mix the reagent mix with the sample using the electromagnet.

[0263]

[0252] In some embodiments, the analyzer includes a single electromagnet installed in the analyzer at a position near a top edge or portion (or any other directional portion) of the mixing chamber when the test cartridge is installed in the analyzer. After the sample and reagent is loaded into the mixing chamber and the test cartridge is inserted into the analyzer, the magnetic beads are moved through the mixing chamber and the reagent and sample (for example, pulled towards the top of the mixing chamber by the magnetic field of the electromagnet). This mixing action will speed up reagent rehydration and mixing with the sample mechanically.

[0264]

[0253] In some embodiments, the analyzer comprises two or more electromagnets, one located near the top of the mixing chamber when the test cartridge is inserted into the analyzer and a second located near a bottom (or other directional portion that is opposite the top portion) of the mixing chamber when the test cartridge is inserted into the analyzer. The electromagnets may have a fixed position or moving / movable positions and / or orientations relative to each other and the mixing chamber. When the test cartridge is inserted into the analyzer, the sample and reagent mixture is delivered to the reaction wells of the test cartridge. At the same time, a mechanism can be triggered to switch the position of the electromagnets (or otherwise change the field affecting the magnet beads or other mixing objects) to manipulate the magnetic field (or other field) in a way that induces magnetic bead (or mixing object) movement and enhanced mixing. In some embodiments, the electromagnet(s) is controllable such that the magnetic field from each electromagnet can be turned off / on. In embodiments where there is only a single electromagnet or permanent magnet, the magnetic field may not be controllable. In some embodiments, various parameters of the field or force applied to the mixing object can be varied or controlled by the user to control the movement of the mixing object to control the mixing of the reagent and the sample. Such control may be provided by a control circuit or similar component.

[0265] Reducing Inhibitor Impact

[0266]

[0254] As noted above, the sample is mixed with the reagent to perform the testing. For example, the analyzer and test cartridges enable detection of nucleic acid targets in human biological samples or tissues without nucleic acid purification. In some embodiments, the biological samples or tissues include inhibitors that reduce efficacy or capabilities of the testing of the sample with the reagent. For example, human samples such as nasal fluid and saliva contain one or more of lactoferrin, lysozyme, RNases, DNases, or other nuclease, which can either inhibit reverse transcriptase enzymes or degrade viral target RNA. Thus, the inhibitors in the samples can reduce the detection sensitivity of the test.

[0267]

[0255] By reducing the effects or impacts of the inhibitors in the samples, the tests can provide improved detection sensitivities. In some embodiments, antibodies, proteinase, Aptamers, high affinity competitive binding proteins, or other agents (hereinafter antibodies) are added to the sample (for example, in the mixing chamber) by including the antibodies in the reagent or by introducing the antibodies into the mixing chamber independently. Thus, the antibodies can be added directly to the sample and reagent buffer. In some embodiments, activity of proteinase, such as proteinase K, will be inactivated by a chemical activated heater.

[0256] In particular, certain additives may improve assays for detecting nucleic acids in samples, for example cloacal samples. Cartridges in accordance with the present disclosure may include bovine serum albumin (BSA). In some examples, the cartridge may include BSA as a dried reagent that is rehydrated during operation of the cartridge. Suitable amounts of BSA to include in dried reagents are discussed herein. In some examples, the dried reagents may comprise 0.15% to 0.45% BSA by weight. In some examples, the dried reagent may comprise about 0.30% BSA. Without being bound to a particular theory, it is believed that BSA acts as a stabilizer to the sample.

[0268]

[0257] It may be desirable to include a non- denaturing detergent in a sample buffer to reduce inhibition. In some examples, sample buffers can include octylphenoxy poly(ethyleneoxy)ethanol. Such sample buffers may be particularly useful for cloacal samples. Cartridges, sample filtration devices, and methods in accordance with the present disclosure may all include sample buffers including octylphenoxy poly(ethyleneoxy)ethanol. Such sample buffers may include octylphenoxy poly(ethyleneoxy)ethanol at concentrations of 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, 2.0%, 2.1%, 2.2%, 2.3%, 2.4%, 2.5%, 2.6%, 2.7%, 2.8%, 2.9%, 3.0%, 3.1%, 3.2%, 3.3%, 3.4%, 3.5%, 3.6%, 3.7%, 3.8%, 3.9%, 4.0%, 4.1%, 4.2%, 4.3%, 4.4%, 4.5%, 4.6%, 4.7%, 4.8%, 4.9%, or 5.0% by volume, or within a range defined by any two of these values. In some examples, the sample buffer may include octylphenoxy poly(ethyleneoxy)ethanol at a concentration 0.1% to 5% by volume. In some examples, the sample buffer may include octylphenoxy poly(ethyleneoxy)ethanol at a concentration 0.1% to 3% by volume. In some examples, the sample buffer may include octylphenoxy poly(ethyleneoxy)ethanol at a concentration of 1% by volume. Without being bound to a particular theory, it is believed that octylphenoxy poly(ethyleneoxy)ethanol acts to limit inhibitors of LAMP present in biological samples, particularly cloacal samples.

[0269]

[0258] Alternatively, or additionally, one or more of the swab assembly insertion point, the transition point and a fluid path of the test cartridge, and the mixing chamber are coated with the antibodies, allowing the sample and the reagent to mix with the antibodies and reduce the impact of the inhibitors in the sample as the sample flows through the test cartridge and into the mixing chamber. Thus, by coating the cartridge sample collection port and / or the wall of the fluid channels with antibodies, during the process of sample collection and delivery, these components of the sample will interact and bind to the antibodies. Therefore, the inhibitors can be removed from the reaction of the viral target amplification and detection. In some embodiments, one or more of the cartridge sample collection port, the mixing chamber, a test well, and the fluid channels includes an antibody (e.g., a SingleChain Fragment Variable (“scFv”)) that reduces effects of inhibitors that exist in the sample (for example, as a coating on a wall or similar surface relative to the cartridge sample collection port, the mixing chamber, a test well, and the fluid channels or injected into or otherwise released into the sample, reagent, or sample / reagent mixture).

[0270]

[0259] Other approaches may be suitable for reducing the impact of inhibitors. For example, sample filtration through resin beads, through graphene, and / or through activated charcoal prior to analysis may remove at least some inhibitors from a biological sample. Addition of polysorbate 20, GuHCL, sodium azide, betaine, and / or dithiothreitol (DTT) to sample buffer may reduce activity of some inhibitors present in a biological sample. Additionally or alternatively, a swab having a sample can be prewashed, prior to the addition of an elution buffer, to remove some inhibitors. As an example, EDTA may be used to prewash a swab. Use of commercial kits to remove amplification inhibitors prior to introducing a sample to the cartridge may also be desirable. Such commercial kits can remove at least some inhibitors (e.g., polyphenolics, humic / fulvic acids, tannins, melanin) from certain samples.

[0271] Sample Deposit

[0272]

[0260] Certain biological samples may include various components that may affect efficacy of the test performed on the sample. For example, nasal fluid or saliva samples from humans contain high concentration of inorganic salts, antimicrobial enzyme lysozymes, immunoglobulins, and glycoproteins such as lactoferrin and mucins. The amplification described herein may be RT-LAMP that detects the viral RNA or DNA in the sample. The inorganic salts, such as NaCl, in the sample will affect or interact with the salt concentration of the reagent and the Tm (melting temperature, the temperature at which an oligonucleotide is 50% annealed to its template) of the RT-LAMP assay primers. Thus, the inorganic salts may cause nonspecific amplification issues based on these interactions. The RNase in the sample may cause viral RNA target degradation, and other proteins in the sample may inhibit the enzyme activity for downstream target detection, as described above.

[0261] In many instances, a pipette or a swab may be used to deposit the sample into the test cartridge. The pipette and swab may be designed to minimize any effects on the sample. A replacement sample insertion device may be used to actively effect the sample, more specifically, to remove the salt and proteins in the sample, and generate a “purified” sample. The sample insertion device may load the purified sample directly into the test cartridge so that the purified sample can be tested as described herein without effect from the salts and other proteins.

[0273] Example Methods of Sample Collection and Testing

[0274]

[0262] FIGs. 11A-11F illustrate methods of collecting and using samples with cartridges in accordance with the present disclosure. It is to be understood that a method in accordance with the present disclosure may use any of the steps discussed herein. Sample collection and filtration devices to be used with the cartridge in the context of methods of FIGs. 11 A-l IF are discussed herein.

[0275]

[0263] FIG. 11A illustrates a flowchart of a method 1100 of collecting samples and testing the collected samples using, for example, the cartridge 100 and the analyzer 300 described herein. At block 1102, a swab is inserted into the sample receptacle 142. For example, the swab may be inserted into the sample receptacle 142 until the bristles or flock of the swab moves past the scraper 150 and towards the bottom of the sample receptacle 142. At block 1104, the swab is broken or cut while positioned inside the sample receptacle 142. Once broken or cut, a portion of the swab with the bristles or flock (for example, a portion with mucus or other types of samples) can remain in the sample receptacle 142 and a remaining portion of the swab (for example, a portion without mucus or other types of samples) may be discarded optionally at block 1106. At block 1108, the cap 110 of the sample receptacle 142 is closed. Once the cap 110 is closed, the portion of the swab with mucus or other types of samples is retained inside the sample receptacle 142. At block 1110, the cartridge 100 is inserted into the analyzer 300 for testing.

[0276]

[0264] FIG. 11B illustrates a flowchart of a method 1130 of collecting samples and testing the collected samples using, for example, the cartridge 100 and the analyzer 300 described herein. At block 1132, a swab is inserted into the sample receptacle 142 of the cartridge 100. As described herein, the swab may be inserted into the sample receptacle 142 past the scraper 150 (for example, moving beyond the scraper 150 towards the bottom portion of the swab receptacle). At block 1134, the swab is fixed in position inside the sample receptacle 142. For example, the sample receptacle 142 includes the retainer 180 as described herein that can hold the swab in place while positioned within the sample receptacle 142. While the swab is held in place within the sample receptacle 142, the cartridge 100 can be inserted into the analyzer 300. At block 1136, the cartridge 100 is inserted into the analyzer 300 for testing.

[0277]

[0265] FIG. 11C illustrates a flowchart of a method 1150 of collecting samples and testing the collected samples using, for example, the cartridge 100 and the analyzer 300 described herein. At block 1152, the integrated rupture feature (for example, a blister) 144 of the cartridge 100 is pressed. Pressing the integrated rupture feature 144 can rupture a membrane between the integrated rupture feature 144 and the reagent blister 140, and cause solution (or solutions) previously stored inside the integrated rupture blister 144 to be released into the sample receptacle 142. The integrated rupture feature 144 can store a buffer and / or a reagent needed for testing the collected samples. At block 1154, a swab is inserted into the sample receptacle 142 of the cartridge 100. At block 1156, the swab, while positioned inside the sample receptacle 142, is twisted or rotated once or a plurality of times e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 twists or revolutions. This can help the liberation of mucus or other types of samples from, for example, the bristles or flock of the swab. In some examples, the swab is twisted or rotated five times. At block 1158, the swab is removed from the sample receptacle 142. At block 1160, the sample receptacle 142 is sealed by closing the cap 110. Closing the cap 110 ensures that the collected sample does not leak out from the sample receptacle 142. At block 1162, the reagent blister 140 is pressed. Pressing the reagent blister 140 can facilitate solution stored inside the reagent blister 140 (which can include both the solution previously stored inside the integrated rupture blister and the solution previously stored inside the reagent blister) to towards, for example, the sample mixing and macrofiltration region 130 of the cartridge 100.

[0278]

[0266] FIG. 1 ID illustrates a flowchart of a method 1170 of collecting samples and testing the collected samples using, for example, the cartridge 100 and the analyzer 300 described herein. At block 1172, elute the swab (e.g., the swab 1200) into a filtration device. Such a filtration device may be a buffer vial (e.g., the buffer vial 1300), a vial adaptor (e.g., the vial adapter 1400), or a pipette that can couple with a filter adaptor (e.g., a filter adaptor 2302 or 2402). At block 1174, filter the sample. For a buffer vial or a vial adaptor, this step may include actuating a moving part of the buffer vial or vial adaptor to cause fluid sample to flow through a filter. For a filter adaptor, this step may include using a pipette to flow liquid sample through a filter of the filter adaptor. At block 1176, transfer the filtered sample to the cartridge. When using the filter adaptor or the vial adaptor in accordance with the present disclosure, the filter adaptor or vial adaptor may be coupled to the cartridge such that filtered sample is transferred to the cartridge as the sample is filtered. When using a buffer vial, a user may transfer filtered sample fluid from the buffer vial to a sample receptacle of the cartridge. At block 1178, insert the cartridge into an analyze (e.g., analyzer 300). The step of inserting the cartridge into the analyzer may cause rupture of one or more blisters of the cartridge, which in turn can cause buffer fluid and / or sample fluid to flow through fluidic channels of the cartridge.

[0279]

[0267] FIG. 1 IE illustrates a flowchart of a method 1180 of collecting samples and testing the collected samples and using, for example, the cartridge 100 and the analyzer 300 described herein. At block 1182, insert a swab (e.g., the swab 1200) into a sample receptacle (e.g., sample receptacle 142) of the cartridge. The swab head may optionally be broken off within the sample receptacle such that a sample receptacle cap can be closed to cover the sample receptacle. At block 1184, elute the sample from the swab. In some examples, buffer for eluting the sample from the swab can be introduced from a buffer chamber of the cartridge. In some such examples, insertion of the cartridge into an analyzer may cause a reagent blister of the cartridge to rupture, releasing the buffer. In other examples, elution may be introduced to the sample receptacle by the user, for example via a pipette. At block 1186, filter the sample. The eluted sample may flow through a filter of the cartridge (e.g. filters 204 or 208). At block 1188, analyze the sample. After filtration, reagents may be introduced in accordance with the present disclosure for any suitable test assay to be conducted by the cartridge and analyzer. The cartridge can cause the sample to flow to a test well, which may include sensors for sensing a test result of the assay.

[0280]

[0268] FIG. 1 IF illustrates a flowchart of a method 1190. At block 1192, introduce buffer to the sample. The buffer may include a non-denaturing reagent, for example octylphenoxy poly(ethyleneoxy)ethanol. At step 1194, filter the sample. Filtration may remove solid particles that may interfere with functioning of a cartridge in accordance with the present disclosure. At step 1196, mix the filtered sample with dried reagents. The dried reagents may include reagents for carrying out an assay. The dried reagents may include additives for improving stability of the sample (e.g., BSA). At step 1198, analyze the sample. Analysis may include any suitable assay discussed herein (e.g., LAMP).

[0281] Swab

[0282]

[0269] Aspects of the present disclosure relate to a swab for collecting cloacal samples. FIG. 12 shows an example swab 1200, which includes a swab head 1202, a handle 1204, a shaft 1208, optionally a stability disc 1210, a tip 1214, and optionally a distal shaft portion 1212. Such a swab 1200 may be used with any of the sample collection, elution, or cartridge devices discussed herein.

[0283]

[0270] The swab head 1202 may include a porous material that can uptake sample material, for example cloacal matrix. The size and dimensions of the swab head 1202 may allow for gathering a cloacal sample, for example from poultry. As such, the swab head 1202 may be conical in shape. A length of the swab head 1202 may be larger than a width of the swab head 1202. In some examples, the swab head 1202 can be inserted into a sample receptable of a cartridge of the present disclosure (for example, the sample receptacle 142 of the cartridge 100). Sample may be eluted off of the swab head 1202 while the swab head 1202 is positioned within a sample receptacle. In other examples, the sample may be eluted off the swab head 1202 into a separate container (e.g., a buffer vial, vial adaptor, or filter adaptor in accordance with the present disclosure) before the sample is introduced to the sample receptacle. The shaft point 1208 may include a stability disc 1210 positioned within the swab head 1202. The stability disc 1210 can act to inhibit motion of the swab head 1202 relative to the shaft 1208. Additionally or alternatively, the stability disc 1210 can provide structure against which a scraper of the cartridge (e.g., scraper 150) can push against to express fluid sample from the swab head 1202.

[0284]

[0271] Before elution, the swab head 1202 can be inserted into a formed membrane. The formed membrane may cover the outer surface of the swab head 1202. The formed membrane can trap solid particles having a size larger than the pore size of the membrane from being eluted. The formed membrane can include pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The formed membrane can include pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the pore sizes of the formed membrane are from 20 to 50 pm in diameter.

[0285] Buffer Vial

[0286]

[0272] Aspects of the present disclosure relate to a buffer vial that can filter solid particles from target molecules (e.g., target nucleic acids). FIGs. 13A and 13B show an example of such a buffer vial 1300. The buffer vial 1300 can include a tube 1302, a cap 1304, a filter 1306, and a rod 1308. The cap 1304 can reversibly couple to an open end of the tube 1302, for example by a threaded connection. The filter 1306 is attached to the rod 1308. The rod 1308 is slidingly coupled to the cap 1304. When the cap 1304 is attached to the tube 1302, a user can push the rod 1308 to move the filter 1306 from close to the cap 1304 to a closed end 1314 of the tube 1302. A user can introduce a fluid sample to the tube 1302. In some examples, the fluid sample may be introduced to the tube 1302 by eluting the sample off a swab (e.g., the swab 1200). Once the sample is eluted to the tube 1302, the filter 1306 can be placed in the tube 1302 and the cap 1304 can be coupled to the tube 1302 as shown in FIG. 13 A.

[0287]

[0273] FIG. 13B shows the buffer vial 1300 when the filter 1306 is at an intermediate distance between the cap 1304 and the closed end 1314. In a first portion 1310 of the tube 1302, the solid particles have been removed due to movement of the filter 1306. The solid particles are restricted to a second portion 1312 of the tube 1302. The filter 1306 and the interior of the tube 1302 may mutually impart frictional force sufficient to hold the filter 1306 in place relative to the tube 1302 even if the cap 1304 is removed from the tube 1302. Removal of the cap 1304 from the tube 1302 allows a user to remove sample fluid containing no or minimal solid particles from the first portion 1310 for analysis, for example by a cartridge in accordance with the present disclosure. As an illustrative example, sample fluid may be removed from the first portion 1310 by pipette.

[0288]

[0274] The filter 1306 can include pores of suitable size to filter out solid particles from a fluid sample while allowing passage of target molecules (e.g., nucleic acids) from the fluid sample. The filter 1306 can include pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The filter 1306 can include pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the pore sizes are from 20 to 50 pm in diameter. Vial Adaptor

[0289]

[0275] Aspects of the disclosure herein concern an adapter that facilitates the flow of liquid constituents, for example a liquid sample, from a standard vial to a target sensing device or a diagnostic device e.g., a cartridge configured to detect a pathogen. The adapter may be coupled to a standard vial and a standard target sensing device without any modification to the vial or the target sensing device. In some aspects, the adapter inhibits or prevents re-aspiration of the liquid constituent into the vial. In some aspects, the adapter comprises or serves as a filter for the liquid constituent.

[0290]

[0276] FIGs. 14A and 14B illustrate various views of an example vial adapter 1400. The vial 1402 can be, in some embodiments, an off-the-shelf vial or the vial can be specifically or custom manufactured. The vial adapter 1400 can include a cap 1410 and a piston 1450. The cap 1410 can be removably coupled to a vial 1402 during use and can be decoupled after use. For example, the vial 1402 can include a threaded or interlocking portion 1510 (an example is shown in FIG. 15), which corresponds to a threaded or interlocking portion 1920 (an example is shown in FIG. 19B) of the cap 1410. The piston 1450 can be slidingly inserted into a cavity formed by the cap 1410 and translate towards the vial 1402 through the cap 1410. In some embodiments, the cap 1410 and piston 1450 can include a mechanism that allows the piston 1450 to translate when it is rotated relative to the cap 1410. The translational movement of the piston 1450 (for example, distal translation through the cap 1410 towards the vial 1402) can generate a positive pressure and / or cause fluid stored in the vial 1402 to flow out of the vial 1402 through the vial adapter 1400.

[0291]

[0277] The piston 1450 can include a proximal end (for example, an end opposite the vial 1402 as shown in FIG. 14A) that can removably couple with an opening of a target sensing device or a diagnostic device (for example, see FIGs. 21 A and 21B).

[0292]

[0278] FIG. 15 illustrates an exploded view of the vial adapter 1400 and the vial 1402. The cap 1410 can include a distal end 1514 (for example, an end that is proximate to the vial 1402) and a proximal end 1516 (for example, an end that is distal from the vial 1402). The proximal end 1516 can include an opening dimensioned to receive the piston 1450. The distal end 1514 can include an opening dimensioned to couple with the vial 1402. For example, the distal end 1514 of the cap 1410 can include an inner surface having the threaded portion 1920 (see FIG. 19B). Additionally, the distal end 1514 of the cap 1410 can extend over at least a portion of the threaded portion 1510 of the vial 1402 when coupled to the vial 1402. The coupling between the vial 1402 and the cap 1410 can be leak-proof to prevent or inhibit fluid stored in the vial 1402 from leaking during the transfer process. The cap 1410 can include one or more wings 1512. The wings 1512 may assist a user in tightening the cap 1410 onto and / or removing the cap 1410 from the threaded portion 1510 of the vial 1402. The cap 1410 can include furrows 1518. The furrows 1518 can help a user grip and / or rotate the cap 1410.

[0293]

[0279] The vial 1402 can include the threaded portion 1510 and a protrusion 1532. The protrusion 1532 can engage the distal end 1514 of the cap 1410 and stop further distal translation of the cap 1410.

[0294]

[0280] The piston 1450 can include a seal 1502 and a piston body 1520. The seal 1502 and the piston body 1520 can be removably coupled. The seal 1502 can be positioned on and / or within an indentation 1504 of the piston body 1520. The seal 1502 can be dimensioned such that its outer circumference 1506 can abut against an inner surface (for example, inner surface 2004 shown in FIG. 20A) of the vial 1402 to create a watertight barrier. The watertight barrier between the inner surface 2004 of the vial 1402 and the outer circumference 1506 of the seal 1502 can be maintained while the piston 1450 is distally translated through the cap 1410 towards the vial 1402. Additionally, the watertight barrier between the inner surface 2004 of the vial 1402 and the outer circumference 1506 of the seal 1502 can prevent or inhibit fluid stored in the vial 1402 from leaking between the seal 1502 1500 and the inner surface 2004 of the vial 1402 during use. Additionally, the barrier between the inner surface 2004 of the vial 1402 and the outer circumference 1506 of the seal 1502 can generate a positive pressure gradient or volume displacement in the vial 1402, which can cause fluid stored in the vial 1402 to flow out of the vial 1402 via the vial adapter 1400.

[0295]

[0281] In some embodiments, the seal 1502 can be made from a material that can generate desired amount of friction against the inner surface 2004 of the vial 1402 (for example, inner surface 2004 shown in FIG. 20A). In some embodiments, the seal 1502 can include an elastomer.

[0296]

[0282] The piston body 1520 of the piston 1450 can include a head 1522, a groove 1528, a stopper 1530, and a proximal portion 1540. The head 1522 can include a distal end 1524 and a filter 1508. The distal end 1524 can be shaped such that when the head 1522 is inserted into the vial 1402, the distal end 1524 matches the interior surface of the vial 1402. Conformance between the head 1522 and the interior surface of the vial 1402 can aid in causing all and / or substantially all of the fluid stored in the vial 1402 to flow through a channel 1806 (as shown in FIGs. 18A and 18B) of the piston body 1520 and the filter 1508 when the piston 1450 is actuated towards the vial 1402.

[0297]

[0283] The groove 1528 can be formed on the piston body 1520. In some embodiments, there are one, two, three, four, five or six grooves 1528 formed on the piston body 1520, though other numbers of grooves 1528 may be suitably implemented. The embodiment depicted in FIGS. 14A-21B includes two grooves 1528. The groove 1528 can be formed in the piston body 1520 to follow a spiral, helical, and / or screw-like path, running along at least a portion of the length of the piston body 1520 while running along at least a portion of the outer circumference of the piston body 1520. The groove 1528 can engage a detent of the cap 1410 (for example, detent 1910 shown in FIGs. 19A and 19B). When engaged, the groove 1528 and the detent can cause the piston 1450 to translate relative to the cap 1410 when the piston 1450 rotates relative to the cap 1410. The groove 1528 can include locks 1526a and / or 1526b that can engage the detent (for example, detent 1910 shown in FIGs. 19A and 19B) of the cap 1410 to at least partially restrict the movement of the piston 1450 through the cap 1410 and / orto provide tactile feedback. Each oflocks 1526a and 1526b may include one or more ridges that can slightly or substantially inhibit motion of a detent of the cap 1410 past the lock 1526a and / or 1526b. To twist the piston 1450 such that the detent 1910 travels beyond the lock 1526a and / or 1526b, the user can exert a threshold force. The threshold force may be larger than a force ordinarily needed to twist the piston 1450 when the lock 1526a and / or 1526b is not engaged by the detent 1910 of the cap 1410. In some embodiments, the piston 1450 includes a lock 1526a corresponding to a pre-actuation position of the piston 1450 relative to the cap 1410. In such embodiments, the user can exert at least the threshold force on the piston 1450 to move the detent past the lock 1526a to initiate actuation of the piston 1450. The lock 1526a may provide a user with tactile feedback that actuation of the piston 1450 has begun. In some embodiments, the piston 1450 includes a lock 1526b corresponding to a post-actuation position of the piston 1450 relative to the cap 1410. In such embodiments, the user can exert at least the threshold force on the piston 1450 to move the detent past the lock 1526b to complete actuation. The lock 1526b may provide a user with tactile feedback that the actuation of the piston 1450 is complete. In some embodiments, there may be additional locks along the groove 1528. Such additional locks may provide tactile feedback to a user that a particular intermediate volume of fluid has been deployed from the vial 1402.

[0298]

[0284] The stopper 1530 can extend radially outward from an outer circumference of the piston body 1520 proximate to the proximal portion 1540. The stopper 1530 can engage the proximal end 1516 of the cap 1410 to prevent further movement (for example, distal translation) of the piston 1450 with respect to the cap 1410 (for example, through the cap 1410). Additionally, the stopper 1530 can abut against an opening of a testing device (e.g., a cartridge for sensing and / or identifying pathogens, genomic materials, proteins, and / or other small molecules or biomarkers) or another storage device (for example, for fluid samples) during use such that fluid stored in the vial 1402 can be transferred to the testing device or the other storage device via the vial adapter 1400.

[0299]

[0285] In some embodiments, the piston 1450 and the cap 1410 may be integrated into a single device, with the piston 1450 being able to move (for example, translate distally or proximally and / or rotate) with respect to the cap 1410.

[0300]

[0286] FIG. 16 illustrates a perspective view of the vial adapter 1400. The piston 1450 can include a distal aperture 1602 (shown in dashed lines) formed on a distal end 1524 of the head 1522 and covered by the filter 1508. During use, as the piston 1450 distally translates towards the vial 1402, the head 1522 can move, for example, into a cavity 2002 (see FIGs. 20A-20C) of the vial 1402. As the head 1522 moves into the cavity (for example, the cavity 2002) of the vial 1402, it displaces the volume of the cavity and generates a positive pressure change in the vial 1402. Coupled with the water-tight barrier between the seal 1502 and the inner surface (for example, the inner surface 2004) of the vial 1402, the distal movement of the piston 1450 can cause flow of fluid stored in the vial 1402 via the filter 1508 and the distal aperture 1602 and out of the vial 1402 via the vial adapter 1400. In some embodiments, at least a portion of the head 1522 extends into a portion of the vial 1402. The proximal portion 1540 can include one or more wings 1604 that stopper 1530. The one or more wings 1604 can allow a user to more easily twist the vial adapter 1400.

[0301]

[0287] With references to FIGs. 17A-18B, the seal 1502 can be removably coupled with head 1522 of the piston body 1520. The seal 1502 may be an o-ring. The shape and dimensions of the indentation 1504 can correspond to the shape of seal 1502. The indentation 1504 can prevent the seal 1502 from slipping off the head 1522 of the piston body 1520. The filter 1508 is attached to the head 1522 so as to resist detachment due to fluid flow through the filter 1508 and the distal aperture 1602. In some embodiments, the filter 1508 is attached to the head 1522 via a heat stake bond. The filter 1508 can include pores of suitable size to filter out solid particles from a sample while allowing passage of target biological material (e.g., nucleic acids). The filter 1508 can include plug pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The filter 1508 can include plug pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the plug pore sizes are from 20 to 50 pm in diameter.

[0302]

[0288] With reference to FIGs. 18A-18B, the piston body 1520 can include a distal opening 1602, a filter 1508 covering the distal opening 1602, a proximal opening 1804, and a channel 1806 extending and formed between the distal opening 1602 and the proximal opening 1804. FIG. 18A presents an exploded cross-sectional view of the piston body 1520. FIG. 18B is a non-exploded cross-sectional view of the piston body 1520.

[0303]

[0289] In some embodiments, the width of the channel 1806 can be uniform along the length of the piston 1450 or vary along the length of the piston 1450. For example, the width of the channel 1806 can, as shown in FIGs. 18A and 18B, gradually increase from the distal opening 1602 to the proximal opening 1804. In another example, the width of the channel can remain substantially the same between the majority of the channel 1806 between the distal opening 1602 and the stopper 1530, and increase significantly at the proximal portion 1540. The gradual increase of the width of the channel 1806 can facilitate flow through the channel 1806.

[0304]

[0290] As described herein and shown in FIG. 18 A, the piston body 1520 can include grooves 1528. The grooves 1528, when engaged with detents of the cap 1410, can cause the piston 1450 to translate relative to the cap 1410 due to rotational motion relative to the cap 1410. The grooves 1528, when engaged with detents of the cap 1410, can cause the piston 1450 to rotate relative to the cap 1410 due to translational motion of the piston 1450 relative to the cap 1410.

[0291] FIGs. 19A and 19B illustrate various views of the cap 1410 of the vial adapter 1400. The cap 1410 can include a proximal opening 1906, a distal opening 1902, an inner circumference 1904, detents 1910, a first end 1514, a threaded portion 1920, and wings 1512. The proximal opening 1906 can be dimensioned to receive the piston 1450. The distal opening 1902 can be dimensioned to, for example, extend over at least a portion of the vial 1402 (for example, the threaded portion 1510 of the vial 1402). The threaded portion 1920 can be formed on an inner surface of the distal opening 1902 and can couple with a corresponding threaded portion (for example, the threaded portion 1510) of the vial 1402. As described herein, the interaction between the threaded or interlocking portion 1920 of cap 1410 and the threaded or interlocking portion 1510 of the vial 1402 can allow the cap 1410 of the vial adapter 1400 to removably couple with the vial 1402.

[0305]

[0292] The detents 1910 can be shaped to engage with grooves of the piston. The detents can be positioned on an inner circumference 1904 of the cap 1410. The detents 1910 may have a conical shape. The detents 1910 may be conical in shape with a rounded or flattened top 1912, as shown in FIGs. 19A and 19B. The cap 1410 may include the same number of detents 1910 as grooves on the piston 1450. In embodiments with two detents, the detents 1910 can be arranged opposite each other within the interior surface of the cap arranged (i.e. 180° from each other on the circumference of an interior surface of the cap). In embodiments with three or more detents, each detent 1910 can be arranged equidistant from the proximate detents (i.e. detents are (360 / «)° from each other on the circumference of an interior surface of the cap, where n is the total number of detents).

[0306]

[0293] In some embodiments, the detent 1910 and groove 1528 can each be shaped and / or formed from a material that can generate desired amount of friction when the detent 1910 and groove 1528 slidingly contact.

[0307]

[0294] FIGs. 20A-20C illustrate cross-sectional views of the vial adapter 1400, showing different positions of the piston 1450 during use and motion of components of the vial adapter 1400 and fluid contained therein. FIG. 20A illustrates relative position of the piston 1450 of the vial adapter 1400 with respect to the vial 1402 and the cap 1410 prior to actuation (for example, distal translation of the piston 1450 towards the vial 1402). Prior to actuation, at least a portion of the piston 1450 (for example, the head 1522) may be positioned inside the vial 1402. Additionally, the outer circumference of the piston 1450 (for example, the outer surface of the seal 1502) can abut against the inner surface of the vial 1402 to provide water-tight barrier between the inner surface of the vial 1402 and the piston 1450. The water-tight barrier can prevent or inhibit flow of fluid stored in the vial 1402 being diverted into a space between the cap 1410 and the piston body 1520 of the piston 1450. In some embodiments, the detents 1910 the cap 1410 can be engaged by locks 1526b within the grooves 1528 of the piston 1450.

[0308]

[0295] FIGs. 20B and 20C illustrate relative position of the piston 1450 with respect to the vial 1402 and the cap 1410 during distal translation of the piston 1450. As described herein, the piston 1450 (or the piston body 1520 of the piston 1450) can be actuated and / or translated distally towards the vial 1402 (for example, as indicated by a directional arrow A). When the piston 1450 moves in the direction A, the head 1522 of the piston 1450 moves further into the vial 1402. This causes a positive pressure change and / or volume displacement in the vial 1402, which causes a flow of fluid stored in the vial 1402 out of the vial 1402 via the filter 1508, the distal opening 1802, the channel 1806, and the proximal opening 1804 (for example, as indicated by directional arrows B).

[0309]

[0296] When the piston 1450 is fully actuated (for example, at its distal-most position with respect to the cap 1410), the stopper 1530 of the piston 1450 can abut against the proximal end 1516 of the cap. Additionally, the detent 1910 may engage the lock 1526b when the piston 1450 is fully actuated. The head 1522 may contact an interior surface 2010 of the cavity 2002.

[0310]

[0297] With reference to FIGs. 21A and 21B, the vial adapter 1400 can be used to transfer, for example, fluid stored in the vial 1402 to a receiving device 2100. The receiving device 2100 may be a cartridge in accordance with the present disclosure. The proximal portion 1540 of the piston 1450 can be inserted into an opening 2102 of the receiving device 2100. To prevent accidental leak from the vial 1402 via the vial adapter 1400, the vial 1402 and the vial adapter 1400 can be oriented to have the vial 1402 positioned below the vial adapter 1400 while the proximal portion 1540 is being coupled to the opening 2102 of the receiving device 2100. In some embodiments, the proximal portion 1540 and the opening 2102 can be coupled via friction fit. In some embodiments, the proximal portion 1540 and the opening 2102 can have corresponding attachment features (for example, threads) that allow the proximal portion 1540 and the opening 2102 to be removably secured to each other.

[0298] Once the vial 1402 and the vial adapter 1400 are coupled to the receiving device 2100, the receiving device 2100 can be positioned below the vial 1402 and the vial adapter 1400. The piston 1450 can then be translated towards the cap 1410 by, for example, rotating the piston 1450 relative to the cap 1410, thereby causing the vial 1402 and the cap 1410 to translate towards the receiving device 2100. Pushing the vial 1402 and the cap 1410 towards the receiving device 2100 can cause the head 1522 of the piston 1450 to move towards or into the vial 1402 to generate positive pressure and / or volume displacement in the vial 1402. The positive pressure and / or volume displacement in the vial 1402 can cause flow of, for example, fluid stored in the vial 1402 to the receiving device 2100.

[0311]

[0299] Other suitable vial adaptors may be used, so long as they include a filter that, upon actuation of the vial adaptor, the sample fluid flows through. For instance, U.S. App. No. 18 / 612911, incorporated herein by reference, describes a vial adaptor including a teeth, rack, and arm mechanism to ensure that the piston of the vial adaptor is actuated in one direction only.

[0312] Method of Using the Vial Adapter

[0313]

[0300] FIG. 22 illustrates an example method 2200 of transferring fluid from a vial (for example, the vial 1402) to a receiving device 2100 (for example, a testing device such as a cartridge in accordance with the present disclosure) using the vial adapter 1400. At step 2202, a vial (for example, the vial 1402) is attached to the cap 1410 of the vial adapter 1400. For example, as described herein, the vial 1402 can include a threaded portion 1510 that can be screwed into the threaded portion 1920 of the cap 1410.

[0314]

[0301] At step 2204, an opening of a receiving device 1700 (for example, a testing device or a cartridge) is coupled to the proximal portion 1540 of the piston 1450. In some embodiments, the opening 1702 of the receiving device 1700 and the proximal portion 1540 can have corresponding coupling features (for example, threads, snap-fit, or a single thread and lock coupling) that allow the piston 1450 to be removably secured to the opening 1702 of the receiving device 1700. In some embodiments, the proximal portion 1540 of the piston 1450 can be coupled to the opening 1702 of the receiving device 1700 via friction fit. At step 2206, the piston 1450 is actuated. As described herein, the piston 1450 may be actuated by rotating the piston 1450 relative to the cap 1410 and vial 1402, thereby distally translating the piston 1450 towards the vial 1402 due to the interaction of the detents 1910 of the cap 1410 and the grooves 1528 of the piston body 1520. When the piston 1450 moves towards the vial 1402, the movements of the head 1522 and the seal 1502 along the inner surface of the vial 1402 can generate positive pressure and / or volume displacement in the vial 1402, which can in turn generate flow out of the vial 1402, through the piston 1450 and into the receiving device 1700.

[0315] Filter Adaptor

[0316]

[0302] In certain aspects, the present disclosure provides for a filter adaptor for filtering a fluid sample as it is introduced to a cartridge by a pipette. The filter adaptor may be compatible with a cartridge in accordance with the present disclosure.

[0317]

[0303] FIG. 23A-23C depict an example filter adaptor 2302. The filter adaptor 2302 can couple at a first end with a pipette 2304 and, at a second end of the filter adaptor 2302, with a cartridge 2306. In particular, the second end of the filter adaptor 2302 can couple with a sample receptacle 2308 of the 2306. FIG. 23A shows an exploded arrangement of the filter adaptor 2302, the pipette 2304 and the cartridge 2306. FIG. 23B shows an arrangement where the pipette 2304 is coupled to the first end of the filter adaptor 2302 and the cartridge 2306 is coupled to the second end of the filter adaptor 2302.

[0318]

[0304] FIG. 23 C is an expanded cross-section view of the filter adaptor 2302 when coupled to the sample receptacle 2308 of the cartridge 2306 and the pipette 2304. The filter adaptor 2302 includes a filter 2310. Sample fluid from the pipette 2304 can flow into the first cavity 2312, through the filter 2310, and to the second cavity 2314 before flowing into the sample receptacle 2308 as indicated by the directional arrows in FIG. 23C. The filter 2310 can include pores of suitable size to filter out solid particles from the fluid sample while allowing passage of target biological material (e.g., nucleic acids) from the fluid sample. The filter 2310 can include plug pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The filter 2310 can include plug pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the plug pore sizes are from 20 to 50 pm in diameter.

[0319]

[0305] FIGS. 24A-24B show another example filter adapter 2402. The filter adapter 2402 can couple with a pipette 2404 and a cartridge 2406 as shown in FIG. 24A. FIG. 24B shows an exploded view of the filter adapter 2402. The filter adapter 2402 can include a pipette portion 2408, a cartridge portion 2410, a filter 2412, a filter support 2414, a first fluid opening 2416, a pipete lock 2418, a first threaded portion 2420, an internal cavity 2422, a cartridge protrusion 2424, a second threaded portion 2426, and a second fluid opening 2428. The filter adapter 2402 can couple to a sample receptacle of the cartridge 2406 via the cartridge protrusion 2424, for example by friction fit. The filter adapter 2402 can couple to the pipette 2404 via the pipete lock 2418. The pipete lock 2418 can reversibly couple to the pipete 2404, for example by twist lock.

[0320]

[0306] The filter adapter 2402 may include two portions, the pipette portion 2408 and the cartridge portion 2410, that can be reversibly coupled, for example via the first threaded portion 2420 and the second threaded portion 2426. In other examples, other mechanisms may be suitable for coupling the pipette portion 2408 and the cartridge portion 2410, for example snap fit or friction fit. When coupled, the pipete portion 2408 and the cartridge portion 2410 can secure the filter 2412 and the filter support 2414 within an internal cavity 2422 of the filter adapter 2402. The filter support 2414 can provide support to the filter 2412 to oppose the flow of sample fluid from the first fluid opening 2416 through the filter 2412 to the second fluid opening 2428 when the filter adapter 2402 is in use.

[0321]

[0307] The filter 2412 can include plug pore sizes of 50 pm in diameter or smaller, such as less than 50, 45, 40, 35, 30, 25, or 20 pm in diameter. The filter 2412 can include plug pore sizes of 20 pm in diameter or larger, such as larger than 10, 15, 20, 25, 30, 35, 40, 45, or 50 pm in diameter. In some examples, the plug pore sizes are from 20 to 50 pm in diameter.

[0322]

[0308] In some embodiments, any of the devices described with respect to the Sample Deposit herein can be integrated or otherwise used together. Furthermore, one or more of these Sample Deposit devices may be integrated with one or more of the testing systems described herein (for example, the testing devices, test cartridges, and readers used in conjunction with the systems and methods of tracking infection potential, as described above). The systems and methods of determining wellness scores for users or individuals may include or utilize one or more primers described herein are able to implement or realize the limits of detection (LOD) described herein with reference to two-step incubation and improve tests performed by the testing device using any improvements described with reference to Dry Reagents, Mixing, Reducing Inhibitor Impact, and Sample Deposit. EXAMPLES

[0323]

[0309] Additional examples are disclosed in further detail in the following examples. The examples provided herein are not in any way intended to limit the scope of the claims.

[0324] Example 1. Inclusion of Bovine Serum Albumin in Dry Reagents

[0325]

[0310] FIG. 25 shows time to reaction (TTR) of a MS2 assay for samples including various BSA and clinical cloacal matrix concentrations. The standard BSA concentration included in dry down was 0.15% (designated as “lx BSA”). 2x BSA indicates 0.30% BSA in the dry down, and 3x BSA indicates 0.45% in the dry down. Clinical cloacal matrix was included at 0% (i.e., “No Matrix”), 5% and 10% concentrations in PBS. Increasing BSA concentration to 2x and 3x of standard appear to lower TTR.

[0326] Example 2, Inclusion of Octylphenoxy Polv(ethyleneoxy)ethanol

[0327]

[0311] FIG. 26 shows time to reaction for a MS2 assay for samples including varying concentrations of octylphenoxy poly(ethyleneoxy)ethanol, a non-denaturing detergent. Clinical cloacal matrix was included at a concentration of 20%. The samples tested included either 1%, 2%, 3%, 4%, 5%, or no octylphenoxy poly(ethyleneoxy)ethanol. A TTR of 60 indicated no target nucleotide was detected. Inclusion of octylphenoxy poly(ethyleneoxy)ethanol appears to improve detection of the MS2 assay.

[0328] Example 3, Combination of Bovine Serum Albumin and Octylphenoxy Polviethyleneoxy (ethanol

[0329]

[0312] FIG. 27 shows time to reaction for samples including various combinations of BSA and octylphenoxy poly(ethyleneoxy)ethanol. The test conditions included lx, 2x, and 3x BSA paired with 0%, 1%, and 2% octylphenoxy poly(ethyleneoxy)ethanol (indicated as “OPEE” in the figure). Nucleic targets included three avian flu sequences: H5N2 28-4, H5N2 28-5, and H7Nx 28-12 and one no-template control (NTC). Clinical cloacal matrix was included at 0% for some samples and 15% for other samples. Of the samples that included clinical cloacal matrix, combining BSA and octylphenoxy poly(ethyleneoxy)ethanol appear to show most improvement over the control.

[0330] Example 4, Buffer Volume

[0331]

[0313] FIGs. 28A and 28B show TTR for various elution buffer volumes. Swabs were used to gather the target of the MS2 assay (“Pos”) or a control sample having no target (“Neg”). Buffer was used to elute the sample from the swabs. For buffer not including octylphenoxy poly(ethyleneoxy)ethanol, the elution volumes were 1 mL, 2 mL, 5 mL, and 10 mL, whereas for buffer including 1% octylphenoxy poly(ethyleneoxy)ethanol by volume, the elution volumes were 2 mL, 3 mL, 4 mL, and 5 mL. Eluting with 3-5 mL of the 1% octylphenoxy poly(ethyleneoxy)ethanol buffer appears to give relatively low TTR.

[0332] Example 5, Column Filtration

[0333]

[0314] FIG. 29 shows TTR for samples subjected to gel filtration and / or dilution. A swab in accordance with the present disclosure was used to collect positive or negative control samples for a MS2 assay. The indicated samples were subjected to column filtration using Cytiva PD Minitrap columns with Sephadex G-25 resin. The indicated samples included clinical cloacal matrix at 20% concentration. For the right- most condition (Column, 1 : 5 Column Matrix), the swab was eluted in 1 mL of elution buffer. The elution was further diluted to 20% volume / volume (i.e., 1:5) before filtering using the column.

[0334] Terminology

[0335]

[0315] The methods disclosed herein comprise one or more steps or actions for achieving the described method. The method steps and / or actions may be interchanged with one another without departing from the scope of the claims. In other words, unless a specific order of steps or actions is required for proper operation of the method that is being described, the order and / or use of specific steps and / or actions may be modified without departing from the scope of the claims.

[0336]

[0316] The term “comprising” as used herein is synonymous with “including,” “containing,” or “characterized by,” and is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. By “consisting of’ is meant including, and limited to, whatever follows the phrase “consisting of.” Thus, the phrase “consisting of’ indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of’ is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of’ indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present depending upon whether or not they materially affect the activity or action of the listed elements.

[0337]

[0317] The articles “a” and “an” are used herein to refer to one or to more than one (for example, at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.

[0338]

[0318] The terms “about” or “around” as used herein refer to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much as 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1% to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length.

[0339]

[0319] The above description discloses several methods and materials of the present invention. This invention is susceptible to modifications in the methods and materials, as well as alterations in the fabrication methods and equipment. Such modifications will become apparent to those skilled in the art from a consideration of this disclosure or practice of the invention disclosed herein. Consequently, it is not intended that this invention be limited to the specific embodiments disclosed herein, but that it cover all modifications and alternatives coming within the true scope and spirit of the invention.

[0340]

[0320] All references cited herein, including but not limited to published and unpublished applications, patents, and literature references, are incorporated herein by reference in their entirety and are hereby made a part of this specification. To the extent publications and patents or patent applications incorporated by reference contradict the disclosure contained in the specification, the specification is intended to supersede and / or take precedence over any such contradictory material.

[0341]

[0321] The practice of the present disclosure will employ, unless indicated specifically to the contrary, conventional methods of molecular biology and recombinant DNA techniques within the skill of the art, many of which are described below for the purpose of illustration. Such techniques are explained fully in the literature. See, e.g., Sambrook, et al., Molecular Cloning: A Laboratory Manual (3rd Edition, 2000); DNA Cloning: A Practical Approach, vol. 1 & II (D. Glover, ed.); Oligonucleotide Synthesis (N. Gait, ed., 1984); Oligonucleotide Synthesis: Methods and Applications (P. Herdewijn, ed., 2004); Nucleic Acid Hybridization (B. Hames & S. Higgins, eds., 1985); Nucleic Acid Hybridization: Modern Applications (Buzdin and Lukyanov, eds., 2009); Transcription and Translation (B. Hames & S. Higgins, eds., 1984); Animal Cell Culture (R. Freshney, ed., 1986); Freshney, R.I. (2005) Culture of Animal Cells, a Manual of Basic Technique, 5th Ed. Hoboken NJ, John Wiley & Sons; B. Perbal, A Practical Guide to Molecular Cloning (3rd Edition 2010); Farrell, R., RNA Methodologies: A Laboratory Guide for Isolation and Characterization (3rd Edition 2005).

[0342]

[0322] The terms “function” and “functional” as used herein refer to a biological, enzymatic, or therapeutic function.

[0343]

[0323] The term “isolated” as used herein refers to material that is substantially or essentially free from components that normally accompany it in its native state. For example, an “isolated cell,” as used herein, includes a cell that has been purified from the milieu or organisms in its naturally occurring state, a cell that has been removed from a subject or from a culture, for example, it is not significantly associated with in vivo or in vitro substances.

[0344]

[0324] The terms “nucleic acid” or “nucleic acid molecule” as used herein refers to polynucleotides, such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), oligonucleotides, fragments generated by the polymerase chain reaction (PCR), and fragments generated by any of ligation, scission, endonuclease action, and exonuclease action. Nucleic acid molecules can be composed of monomers that are naturally-occurring nucleotides (such as DNA and RNA), or analogs of naturally-occurring nucleotides (e.g., enantiomeric forms of naturally-occurring nucleotides), or a combination of both. Modified nucleotides can have alterations in sugar moieties and / or in pyrimidine or purine base moieties. Sugar modifications include, for example, replacement of one or more hydroxyl groups with halogens, alkyl groups, amines, and azido groups, or sugars can be functionalized as ethers or esters. Moreover, the entire sugar moiety can be replaced with sterically and electronically similar structures, such as aza-sugars and carbocyclic sugar analogs. Examples of modifications in a base moiety include alkylated purines and pyrimidines, acylated purines or pyrimidines, or other well-known heterocyclic substitutes. Nucleic acid monomers can be linked by phosphodiester bonds or analogs of such linkages. Analogs of phosphodiester linkages include phosphorothioate, phosphorodithioate, phosphoroselenoate, phosphorodiselenoate, phosphoroanilothioate, phosphoranilidate, or phosphoramidate. The term “nucleic acid molecule” also includes so-called “peptide nucleic acids,” which comprise naturally-occurring or modified nucleic acid bases attached to a polyamide backbone. Nucleic acids can be either single stranded or double stranded. “Oligonucleotide” can be used interchangeable with nucleic acid and can refer to either double stranded or single stranded DNA or RNA. A nucleic acid or nucleic acids can be contained in a nucleic acid vector or nucleic acid construct (e.g. plasmid, virus, bacteriophage, cosmid, fosmid, phagemid, bacterial artificial chromosome (BAC), yeast artificial chromosome (YAC), or human artificial chromosome (HAC)) that can be used for amplification and / or expression of the nucleic acid or nucleic acids in various biological systems. Typically, the vector or construct will also contain elements including but not limited to promoters, enhancers, terminators, inducers, ribosome binding sites, translation initiation sites, start codons, stop codons, polyadenylation signals, origins of replication, cloning sites, multiple cloning sites, restriction enzyme sites, epitopes, reporter genes, selection markers, antibiotic selection markers, targeting sequences, peptide purification tags, or accessory genes, or any combination thereof.

[0345]

[0325] A nucleic acid or nucleic acid molecule can comprise one or more sequences encoding different peptides, polypeptides, or proteins. These one or more sequences can be joined in the same nucleic acid or nucleic acid molecule adjacently, or with extra nucleic acids in between, e.g. linkers, repeats or restriction enzyme sites, or any other sequence that is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths. The term “downstream” on a nucleic acid as used herein refers to a sequence being after the 3 ’-end of a previous sequence, on the strand containing the encoding sequence (sense strand) if the nucleic acid is double stranded. The term “upstream” on a nucleic acid as used herein refers to a sequence being before the 5 ’-end of a subsequent sequence, on the strand containing the encoding sequence (sense strand) if the nucleic acid is double stranded. The term “grouped” on a nucleic acid as used herein refers to two or more sequences that occur in proximity either directly or with extra nucleic acids in between, e.g. linkers, repeats, or restriction enzyme sites, or any other sequence that is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths, but generally not with a sequence in between that encodes for a functioning or catalytic polypeptide, protein, or protein domain.

[0346]

[0326] The terms “peptide”, “polypeptide”, and “protein” as used herein refers to macromolecules comprised of amino acids linked by peptide bonds. The numerous functions of peptides, polypeptides, and proteins are known in the art, and include but are not limited to enzymes, structure, transport, defense, hormones, or signaling. Peptides, polypeptides, and proteins are often, but not always, produced biologically by a ribosomal complex using a nucleic acid template, although chemical syntheses are also available. By manipulating the nucleic acid template, peptide, polypeptide, and protein mutations such as substitutions, deletions, truncations, additions, duplications, or fusions of more than one peptide, polypeptide, or protein can be performed. These fusions of more than one peptide, polypeptide, or protein can be joined in the same molecule adjacently, or with extra amino acids in between, e.g. linkers, repeats, epitopes, or tags, or any other sequence that is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, or 300 bases long, or any length in a range defined by any two of the aforementioned lengths.

[0347]

[0327] The term “% w / w” or “% wt / wt” as used herein has its ordinary meaning as understood in light of the specification and refers to a percentage expressed in terms of the weight of the ingredient or agent over the total weight of the composition multiplied by 100. The term “% v / v” or “% vol / vol” as used herein has its ordinary meaning as understood in the light of the specification and refers to a percentage expressed in terms of the liquid volume of the compound, substance, ingredient, or agent over the total liquid volume of the composition multiplied by 100.

[0348]

[0328] The term “loop-mediated isothermal amplification (LAMP)” as used herein has its plain and ordinary meaning as understood in light of the specification and refers to a method of nucleic acid amplification that is performed isothermally, or without the repeating cycles of temperatures as seen in PCR. LAMP offers a robust way to amplify and detect nucleic acid material from samples such as those derived from subjects rapidly and cost- effectively. Methods of LAMP are well established in the art. Amplification is generally done with a DNA template and a strand-displacing DNA polymerase; inclusion of a reverse transcriptase either added to the amplification solution in a one-pot method, or used to prepare complementary DNA (cDNA) prior to the amplification enables the detection of RNA. Amplification is typically carried out at elevated temperatures (but not near-boiling temperatures as seen with PCR) anywhere within a range between 50°C-70°C that is optimized for the thermotolerant Bst DNA polymerase. However, other temperatures and DNA polymerases can be used with minimal optimization required. LAMP involves at least 4 primers designed for a short region of interest of the nucleic acid target (e.g. a pathogenicity island of a pathogenic bacteria genome). According to conventional primer nomenclature, the 4 primers include two inner primers, forward inner primer (FIP) and backward inner primer (BIP), and two outer primers, F3 and B3. Amplification of the target nucleic acid with the 4 primers results in a characteristic stem-loop containing “dumb-bell” shaped template. Progressive strand synthesis with the inner primers results in the formation of large stem-loop structures from this original template. A later improvement to this original LAMP technique involves the inclusion of 2 loop primers, LF and LB, which hybridize to loop regions of the amplicons and serve as additional points of DNA elongation, significantly improving the speed of complete amplification. The LAMP process can be quantified by several different approaches. Traditional methods include colorimetric detection (e.g. observation of solution turbidity due to the formation of insoluble magnesium pyrophosphate, or with DNA-specific colorimetric dyes), electrophoresis, or antibody-based immunoassays. As disclosed herein in embodiments of detection systems and methods of measuring or analyzing modulations of electrical signals (e.g. impedance or capacitance), LAMP amplification may also be electrochemically detected (e.g. magnesium pyrophosphate accumulation, or the buildup of protons as a result of DNA polymerase activity). Additional information about LAMP can be found in Notomi T et al. “Loop- mediated isothermal amplification of DNA” Nucleic Acids Res. (2000); 28(12):e63 and Nagamine K et al. “Accelerated reaction by loop-mediated isothermal amplification using loop primers” Mol. Cell. Probes (2002); 16(3):223-229, each of which is hereby expressly incorporated by reference in its entirety.

Claims

1. WHAT IS CLAIMED IS:

1. A method for identifying a target molecule in a biological sample, the method comprising: filtering the biological sample to remove at least a portion of solid particles, the solid particles present in the sample; introducing a buffer to the sample to create a first mixture, the buffer comprising at least octylphenoxy poly(ethyleneoxy)ethanol; depositing the first mixture into a sample receptacle of a disposable cartridge; inserting the disposable cartridge into a cartridge receptacle of an analyzer device; mixing the first mixture with dried reagents comprising bovine serum albumin (BSA) and thereby generating a second mixture comprising the first mixture and the BSA; conveying at least a portion of the second mixture to at least one testing well, which comprises at least one dried enzyme and / or a detection agent, such as one or more primers, an antibody or a binding fragment of an antibody, e.g., an ScFv, to generate a third mixture; optionally, increasing a temperature of the at least one testing well to a level sufficient for primer extension or amplification; and measuring an electrical characteristic of at least the portion of the third mixture in the at least one testing well, preferably by measuring a change in impedance at the at least one testing well.

2. The method of claim 1, wherein the filtering comprises using a device, which is not integrated with the disposable cartridge.

3. The method of claim 2, wherein the filtering comprises using a column filter.

4. The method of claim 2, wherein the filtering comprises propelling the biological sample through a filter of a vial adaptor, the vial adaptor comprising: a cap member configured to engage the disposable cartridge; and a piston comprising a distal end, a proximal end, a body extending between the distal end and the proximal end, a channel, and a filter positioned in the channel, the channel configured to allow flow of fluid through the piston and between a distalaperture formed on the distal end of the piston and a proximal aperture formed on the proximal end of the piston, the piston configured to translate distally through the cap member and towards the vial, and the filter configured to block passage of solid particles as the fluid flows through the piston; wherein a distal translation of the piston is configured to cause a positive pressure change in the vial and withdrawal of fluid stored in the vial through the channel of the piston.

5. The method of claim 2, wherein the filtering comprises pipetting the sample through a filter of a filter adapter configured to engage the disposable cartridge and a pipette.

6. The method of claim 1, wherein the filtering is performed using a filter, which is internally disposed within the disposable cartridge.

7. The method of claim 6, wherein the filter comprises a sintered porous plug having plug pore sizes of 20-50 pm in diameter, such as 20, 30, 40, or 50 pm in diameter or having a pore size diameter that is within a range of pore size diameters defined by any two of the aforementioned values.

8. The method of any one of claims 1 to 7, comprising rupturing a reagent blister to introduce the buffer to the sample and / or to rehydrate the dried reagents.

9. The method of claim 8, wherein insertion of the disposable cartridge into the cartridge receptacle causes the rupturing of the reagent blister, the generating of the second mixture, and the conveying of at least the portion of the second mixture to the at least one testing well to generate the third mixture.

10. The method of any one of claims 1 to 9, wherein the buffer comprises 0.1% to 5% of the octylphenoxy poly(ethyleneoxy)ethanol by volume.

11. The method of claim 10, wherein the buffer comprises 1 % of octylphenoxy poly(ethyleneoxy)ethanol by volume.

12. The method of any one of claims 1 to 11, wherein the dried reagents comprises 0.15% to 0.45% BSA by weight.

13. The method of claim 12, wherein the dried reagents comprise 0.30% BSA by weight.

14. The method of any one of claims 1 to 13, wherein the biological sample is diluted prior to introduction into the sample receptacle.

15. The method of claim 14, wherein the biological sample is diluted in water.

16. The method of any one of claims 1 to 15, wherein the biological sample comprises solid particles, which are larger than 50 pm in diameter.

17. The method of any one of claims 1 to 16, wherein the buffer is dried and is rehydrated prior to or concomitantly with generation of the first mixture.

18. The method of any one of claims 1 to 17, wherein the biological sample is a cloacal sample.

19. The method of claim 18, wherein the biological sample is an avian cloacal sample.

20. The method of claim 19, wherein the biological sample is a chicken cloacal sample.

21. An assay cartridge for analyzing a biological sample, comprising a target nucleic acid, the assay cartridge comprising: a cartridge body configured to be received by a reader device, the cartridge body comprising: at least one test well comprising an excitation electrode and a sensing electrode, wherein the at least one test well is configured to receive at least a portion of the biological sample comprising the target nucleic acid; a sample introduction area configured to receive a carrier containing the biological sample; and a fluid path fluidically coupling the sample introduction area to the test well, wherein the fluid path comprises a filter configured to inhibit solid particles of the sample from flowing to the test well; at least one reagent receptacle comprising at least one dried reagent, wherein the at least one dried reagent comprises bovine serum albumin (BSA); and a buffer blister configured to be ruptured when the cartridge body is inserted into the reader device, the buffer blister comprising at least octylphenoxy poly(ethyleneoxy)ethanol, and wherein the rupturing of the reagent blister produces a force that mixes the at least one dried reagent with the buffer and with the biological sample to generate a mixture and moves at least a portion of the mixture through the fluid path to the at least one test well.

22. The assay cartridge of claim 21, wherein the assay cartridge is configured to introduce the buffer to the sample.

23. An assay cartridge for analyzing a biological sample, comprising a target nucleic acid, the assay cartridge comprising: a cartridge body configured to be received by a reader device, the cartridge body comprising: at least one test well comprising an excitation electrode and a sensing electrode, wherein the at least one test well is configured to receive at least a portion of the biological sample comprising the target nucleic acid, wherein the biological sample comprises at least octylphenoxy poly(ethyleneoxy)ethanol; a sample introduction area configured to receive a carrier containing the biological sample; and a fluid path fluidically coupling the sample introduction area to the test well, wherein the fluid path comprises a filter configured to inhibit solid particles of the sample from flowing to the test well; at least one reagent receptacle comprising at least one dried reagent, wherein the at least one dried reagent comprises bovine serum albumin (BSA); and a buffer blister configured to be ruptured when the cartridge body is inserted into the reader device, the buffer blister comprising a buffer, and wherein the rupturing of the reagent blister produces a force that mixes the at least one dried reagent with the buffer and with the biological sample to generate a mixture and moves at least a portion of the mixture through the fluid path to the at least one test well.

24. The assay cartridge of any one of claims 21 to 23, wherein the filter comprises plug pore sizes of 50 pm in diameter or smaller, such as less than 50, 40, 30, or 20 pm in diameter.

25. The assay cartridge of any one of claims 21 to 24, wherein the filter comprises plug pore sizes of 20 pm in diameter or larger, such as larger than 20, 30, 40, or 50 pm in diameter.

26. The assay cartridge of any one of claims 21 to 25, wherein the biological sample comprises solid particles, preferably above 50 pm in diameter.-1 OS-27. The assay cartridge of any one of claims 21 to 26, wherein the at least one dried reagent comprises 0.15% to 0.45% BSA by weight.

28. The assay cartridge of claim 27, wherein the at least one dried reagent comprises 0.30% BSA by weight.

29. The assay cartridge of any one of claims 21 to 28, wherein the biological sample is a cloacal sample.

30. The assay cartridge of claim 29, wherein the biological sample is an avian cloacal sample.

31. The assay cartridge of claim 30, wherein the biological sample is a chicken cloacal sample.

32. The assay cartridge of any one of claims 21 to 31 , wherein the carrier comprises: a channel; and a cap configured to hold the carrier containing the biological sample, the cap further configured to mechanically couple to the cartridge body, wherein mechanically coupling the cap to the cartridge body causes compression of a trapped volume of a fluid to drive at least a portion of the sample through the fluid path into the test well, and wherein the cap comprises: a retaining well having an interior diameter larger than an exterior diameter of the channel; and a retaining structure disposed within the retaining well and configured to retain the channel at a position spaced from a side interior wall and a rear interior wall of the retaining well to form at least one air channel fluidically coupled to an inner end of the channel.

33. The assay cartridge of any one of claims 21 to 32, wherein the cartridge body comprises a base and a translucent cover, the translucent cover comprising a planar surface defining one side of at least one of the test well and the fluid path.

34. The assay cartridge of any one of claims 21 to 33, wherein the cartridge body comprises at least a second test well containing an excitation electrode and a sensing electrode, and a second fluid path fluidically coupling the sample introduction area to the second test well, wherein the second test well is configured to contain at least a portion of the sample, comprising the target nucleic acid.

35. The assay cartridge of any one of claims 21 to 34, wherein the cartridge body comprises a base and a printed circuit board (PCB), the PCB comprising a planar surface defining one side of at least one of the test well and the fluid path.

36. The assay cartridge of claim 35, wherein the PCB comprises a heating element configured to heat the test well.

37. The assay cartridge of claim 35 or 36, wherein the PCB comprises the excitation electrode and the sensing electrode.

38. The assay cartridge of any one of claims 35 to 37, wherein the test well is configured to mix a reagent and the sample into a substantially evenly mixed test fluid.

39. The assay cartridge of claim 38, wherein the reagent comprises one or more dried reagents stored within the test well.

40. The assay cartridge of claim 39, wherein the cartridge body comprises a plurality of test wells, and wherein at least a first test well of the plurality of test wells stores a reagent different from a reagent stored in a second test well of the plurality of test wells.

41. The assay cartridge of claim 39 or 40, wherein the cartridge body comprises a plurality of test wells, and wherein at least two test wells of the plurality of test wells store the same reagent.

42. The assay cartridge of any one of claims 21 to 41, wherein the cartridge body further comprises a mixing chamber positioned between the sample introduction area and the test well along the fluid path, the mixing chamber configured to mix a reagent and the biological sample into a substantially evenly mixed test fluid.

43. The assay cartridge of claim 42, wherein the reagent comprises one or more dried reagents stored within the mixing chamber.

44. The assay cartridge of any one of claims 21 to 43, further comprising a first electrode interface including a first contact pad leading to the excitation electrode and a second contact pad leading to the sensing electrode.

45. The assay cartridge of any one of claims 21 to 44, further comprising a gas- permeable, liquid-impermeable vent fluidically coupled to the test well.

46. The assay cartridge of any one of claims 21 to 45, further comprising a machine- readable cartridge identifier printed thereon, the cartridge identifier associated with one or more test protocols.-HO-47. The assay cartridge of any one of claims 21 to 46, wherein the assay cartridge is a disposable single-use assay cartridge.

48. A buffer vial configured to filter a biological sample prior to transferring the biological sample to a receiving device, the buffer vial comprising: a vial; a cap member configured to removably couple with an opening of the vial; and a filter coupled to a rod, the rod slidingly coupled to the cap, wherein the rod is configured to, when the cap member is coupled to the vial, push the filter from a first position by an open end of the vial to a second position closer to a closed end of the vial than the first position, wherein, at the second position, the filter confines solid particles of the sample to a portion of the vial proximate the closed end.

49. The buffer vial of claim 48, wherein the filter comprises plug pore sizes of 50 pm or smaller, such as less than 50, 40, 30, or 20 pm in diameter.

50. The buffer vial of claim 48 or 49, wherein the filter comprises plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter.

51. The buffer vial of any one of claims 48 to 50, wherein the rod is configured to decouple from the cap member when the filter is in the second position.

52. The buffer vial of any one of claims 48 to 51, comprising a filter support configured to inhibit curvature of the filter as the filter is pushed by the rod.

53. The buffer vial of any one of claims 48 to 52, wherein the biological sample is a cloacal sample.

54. The buffer vial of claim 53, wherein the biological sample is an avian cloacal sample.

55. The buffer vial of claim 54, wherein the biological sample is a chicken cloacal sample.

56. The buffer vial of any one of claims 48 to 55, wherein the vial comprises a nondenaturing detergent.

57. The buffer vial of claim 56, wherein the non-denaturing detergent is octylphenoxy poly(ethyleneoxy)ethanol.

58. A vial adapter configured to filter a biological sample, and transfer the biological sample, from a vial to a receiving device, the vial adapter comprising:a cap member configured to removably couple with an opening of a vial; and a piston comprising a distal end, a proximal end, a body extending between the distal end and the proximal end, a channel, and a filter positioned in the channel, the channel configured to allow flow of fluid through the piston and between a distal aperture formed on the distal end of the piston and a proximal aperture formed on the proximal end of the piston, the piston configured to translate distally through the cap member and towards the vial, and the filter configured to block passage of solid particles as the fluid flows through the piston; wherein a distal translation of the piston is configured to cause a positive pressure change in the vial and withdrawal of fluid stored in the vial through the channel of the piston.

59. The vial adapter of claim 58, wherein the filter comprises plug pore sizes of 50 pm or smaller such as less than 50, 40, 30, or 20 pm in diameter.

60. The vial adapter of claim 58 or 59, wherein the filter comprises plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter.

61. The vial adapter of any one of claims 58 to 60, wherein the cap member comprises an arm, wherein the piston comprises a rack, and wherein the arm is configured to engage the rack to allow the piston to translate distally through the cap member towards the vial and prevent the piston to translate proximally through the cap member away from the vial.

62. The vial adapter of claim 61, wherein the arm comprises a tip, and wherein the tip is curved inwards towards an axis parallel to the body of the piston.

63. The vial adapter of claims 61 or 62, wherein the rack comprises a plurality of teeth, wherein each of the plurality of teeth comprises a leading edge and a stopping edge, and wherein the arm is configured to: slide along the leading edges of the plurality of teeth and the piston is translated distally; and abut one of the stopping edges of the plurality of teeth to prevent the piston from translating proximally.

64. The vial adapter of any of claims 58 to 63, wherein the proximal end of the piston is configured to removably couple with a receiving device.

65. The vial adapter of any of claims 58 to 64, wherein the piston comprises a stopper extending radially and circumferentially outwards from the body of the piston, wherein the stopper is configured to abut the cap member when the piston is in its distal-most position with respect to the cap member.

66. The vial adapter of any of claims 58 to 65, wherein a width of the channel increases from the distal aperture to the proximal aperture.

67. The vial adapter of any of claims 58 to 66, further comprising a cover, wherein the cover is configured to cover the distal end of the body of the piston.

68. The vial adapter of claim 67, wherein the cover is a molded elastomer.

69. The vial adapter of claim 67 or 68, wherein the cover is configured to abut an inner surface of the vial to create a seal between the cover and the inner surface of the vial.

70. The vial adapter of any one of claims 58 to 69, wherein the biological sample is a cloacal sample.

71. The vial adapter of claim 70, wherein the biological sample is an avian cloacal sample.

72. The vial adapter of claim 71, wherein the biological sample is a chicken cloacal sample.

73. A filter adapter for filtering a biological sample having solid particles, the filter adapter comprising: a first portion configured to receive a sample collection device; a second portion adapted for a cartridge sample port; and a filter positioned in a fluid path of the filter adapter, the filter being configured to separate solid particles from a biological sample fluid.

74. The filter adapter of claim 73, wherein the filter comprises plug pore sizes of 50 pm or smaller such as less than 50, 40, 30, 20, or 10 pm in diameter.

75. The filter adapter of claim 73 or 74, wherein the filter comprises plug pore sizes of 20 pm or larger, such as larger than 20, 30, 40, or 50 pm in diameter.

76. The filter adapter of any one of claims 73 to 75, wherein the biological sample is a cloacal sample.

77. The filter adapter of claim 76, wherein the biological sample is an avian cloacal sample.

78. The filter adapter of claim 77, wherein the biological sample is a chicken cloacal sample.

79. The filter adapter of any one of claims 73 to 78, the filter adapter further comprising a filter support positioned within the fluid path and downstream the filter, wherein an upstream surface of the filter support contacts the filter.

80. The filter adapter of any one of claims 73 to 76, wherein the first portion and second portion are configured to interface each other via a threaded connection.

81. The filter adapter of any one of claims 73 to 80, wherein the first portion is configured to create a fluidic seal with a first end of a shell of the filter, and the second portion is configured to create a fluidic seal with a second end of a shell of the filter.

82. The filter adapter of any one of claims 73 to 81, wherein the sample collection device is a pipette.

83. The filter adapter of any one of claims 73 to 81, wherein the sample collection device is a syringe.

84. The filter adapter of any one of claims 73 to 81, wherein the sample collection device is an assay cartridge in accordance with any one of claims 21 to 47.

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