Nucleic acid amplification device

Abstract

The present disclosure relates to nucleic acid amplification devices for testing a bodily fluid sample for a biological analyte and their methods of use. Disclosed herein are nucleic acid amplification systems for untrained users or lay persons. The nucleic acid amplification devices have, e.g., a housing with valves, one or more incubation and detection chambers configured to enable the amplification and the detection of nucleic acids.

Description

NUCLEIC ACID AMPLIFICATION DEVICECROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to, and the benefit of, U.S. Provisional Application No. 63,678,013, filed July 31, 2024, the entirety of which is incorporated herein by reference.BACKGROUND

[0002] The demand for at-home SU testing kits is ever- increasing. However, existing kits require the user to take a sample and then mail the sample to a central lab for processing. These home sampling kits generally take days or weeks to receive the results. Thus, a need still exists for a STI testing device which can be performed in the privacy of a person’s home and also provide fast at-home results.SUMMARY

[0003] At a high level, the present disclosure provide for a nucleic acid amplification device that may be used to amplify a nucleic acid, methods of use thereof, and methods of manufacturing a nucleic acid device as disclosed herein.

[0004] In one aspect, the present disclosure relates to nucleic acid amplification device for testing a bodily fluid sample for a biological analyte. A nucleic acid amplification device includes a housing. The housing includes an inlet port for receiving a sample derived from a bodily fluid and an incubation and detection section configured to enable the amplification and the detection of nucleic acids. The incubation and detection section includes a set of incubation chambers, the set including at least two incubation chambers. Each incubation chamber includes a plurality of nucleic acid amplification compositions, each including a primer configured to amplify nucleic acids indicative of a presence of a biological analyte. The incubation and detection section can include one or more detection chambers. The housing includes a plurality of fluid pathways connecting an inlet port with each incubation chamber of the set of incubation chambers. The housing includes a plurality of detection pathways connecting each incubation chamber to the one or more detection chambers. The incubation and detection section includes a valve separating the set of incubation chambers and the one or more detection chambers, the valve operable to open a fluidic pathway between the set of incubation chambers and the one ormore detection chambers. The housing includes a heating arrangement comprising one or more heating elements adjacent the plurality of incubation chambers..

[0005] In one aspect, a method of detecting a biological analyte using an amplification device is presented. The method includes receiving a sample through an inlet port of an amplification device. The method includes flowing the sample to a set of incubation chambers of the amplification device. The incubation chambers include a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a biological analyte. The method includes heating the set of incubation chambers via a heating arrangements of the amplification device. The heating arrangement includes a set of heating elements positioned beneath the set of incubation chambers. The method includes activating a valve of the amplification device. The valve separates the set of incubation chambers from one or more detection chambers. The method includes flowing the sample from the set of incubation chambers to the one or more detection chambers through the valve. The method includes indicating a present of a threshold amount of DNA and / or RNA via the one or more detection chambers.

[0006] In one aspect the present disclosure provides an amplification device for testing a bodily fluid sample for a biological analyte, the device comprising a housing comprising: an inlet port for receiving a sample derived from a bodily fluid; an incubation and detection section configured to enable the amplification and the detection of nucleic acids, the incubation and detection section comprising: a set of incubation chambers, the set including at least two incubation chambers, wherein each incubation chambers includes a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; and one or more detection chambers; a plurality of fluid pathways connecting the inlet port with each of the incubation chambers; a plurality of detection pathways connecting each incubation chambers to the one or more detection chambers; a valve separating at least one of the set of incubation chambers and the one or more detection chambers, an automated valve (e.g., a wax actuator or shape memory alloy (SMA) coupled to the valve, configured to change shape from a first shape to a second shape, thereby actuating the valve from a closed to open configuration); and a heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers, the one or more heating elements configured to provide heat to each incubation chamber of the set of incubationchambers. In some embodiments, the one or more heating elements are positioned beneath the automated valve or automated valve actuator (e.g., the wax actuator and / or SMA) and produce heat to change the wax actuator or SMA from a first shape to a second shape.

[0007] In some embodiments, the housing comprises a wax actuator. In some embodiments, the housing comprises an SMA. In some embodiments, the housing comprises at least two SMAs in a fluidic channel, the valve (or one or more valves) is positioned between the SMAs in the fluidic channel, and the one or more heating elements provide heat to change the at least two SMAs from a first shape to a second shape, thereby compressing and actuating the valve (or one or more valves) from open to closed configuration. In some embodiments, the change of shape is expansion, contraction, or deflection.

[0008] In some embodiments, the housing further comprises an air bladder, optionally two air bladders, stored within the housing, the air bladder configured to receive air from an incubation chamber of the set of incubation chambers.

[0009] In some embodiments, the housing further comprises an air bladder, optionally two air bladders, stored within the housing, the air bladder configured to receive air from an incubation chamber of the set of incubation chambers.

[0010] In some embodiments, the housing further comprises a hydrophobic vent in fluidic communication with the air bladder, the hydrophobic vent configured to release an amount of air stored in the air bladder. In some embodiments, the hydrophobic vent comprises a sintered Porex Cirtek PD103032 (PTFE) frit.

[0011] In some embodiments, the housing further comprises the air bladder is larger (e.g., significantly larger) than the incubation chamber. In some embodiments, the amplification device does not vent outside of the device (e.g., lacks external venting). In some embodiments, the housing further comprises a set of two air bladders, wherein each incubation chamber of the set of incubation chambers is connected to a respective first outlet in fluidic communication with a respective air bladder and a respective second outlet in fluidic communication with a respective detection chamber.

[0012] In some embodiments, the set of incubation chambers are positioned symmetrically across a horizontal axis of the housing.

[0013] In some embodiments, the detection chambers comprise one or more lateral flow assay (LFA) strips. In some embodiments, the one or more LFA strips are vertically oriented with respect to a base of the housing.

[0014] In some embodiments, the housing further comprises a bubble trap positioned at each outlet of a distribution channel connected to the inlet port and the set of incubation chambers.

[0015] In one aspect the present disclosure provides an amplification device for testing a bodily fluid sample for a biological analyte, the device comprising a housing comprising: an inlet port for receiving a sample derived from a bodily fluid; an incubation and detection section configured to enable the amplification and the detection of nucleic acids, the incubation and detection section comprising: a set of incubation chambers, the set including at least two incubation chambers, wherein each incubation chambers includes a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; and one or more detection chambers; a plurality of fluid pathways connecting the inlet port with each of the incubation chambers; a plurality of detection pathways connecting each incubation chambers to the one or more detection chambers; a valve separating the set of incubation chambers and the one or more detection chambers, the valve operable to open a fluidic pathway between the set of incubation chambers and the one or more detection chambers; an air bladder, optionally two air bladders, configured to receive air from an incubation chamber of the set of incubation chambers; and a heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers, the one or more heating elements configured to provide heat to each incubation chamber of the set of incubation chambers.

[0016] In some embodiments, each nucleic acid amplification composition comprises a different primer configured to amplify nucleic acids indicative of a different illness.

[0017] In some embodiments, a primer of a first nucleic acid amplification composition is configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a second nucleic acid amplification composition is configured to amplify nucleic acids of a another one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a third nucleic acid amplification composition is configured toamplify nucleic acids of a further one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

[0018] In one aspect the present disclosure provides a method of detecting a presence of a biological analyte using an amplification device, comprising: receiving a bodily fluid sample through an inlet port of the amplification device as disclosed herein; flowing the sample to the set of incubation chambers of the amplification device from the inlet port fluid pathways, wherein each incubation chamber of the set of incubation chambers comprises a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; heating the set of incubation chambers via a heating arrangement of the amplification device, the heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers; actuating the valve of the amplification device; flowing the sample from the incubation chamber to the detection chamber through the valve; and indicating a presence of a threshold amount of DNA and / or RNA via the one or more detection chambers.

[0019] In one aspect the present disclosure provides a method of manufacturing and using an amplification device as disclosed herein. In some embodiments, the method comprises

[0020] injection molding a housing component out of a thermoplastic to form: an inlet port; a set of incubation chambers; and a plurality of detection chambers; fluid pathways from the inlet port to the set of incubation chambers; detection pathways from the set of incubation chambers to the one or more detection chambers; inserting a plurality of nucleic acid amplification compositions into each of the incubation chambers, the plurality of nucleic acid amplification compositions each including a primer configured to amplify nucleic acids indicative of a presence of a biological analyte; and mechanically connecting a heating arrangement to a bottom portion of the housing component, the heating arrangement positioned beneath the set of incubation chambers, and optionally valve(s), the heating arrangement comprising a one or more heating elements adjacent the set of incubation chambers and / or valve(s).BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The subject matter of the present disclosure will be explained in more detail in the following text with reference to preferred exemplary and non-limiting embodiments which are illustrated in the attached drawings.

[0022] Fig. 1 illustrates a system having a sample preparation device and an amplification device.

[0023] Figs. 2A-B illustrate perspective views of an amplification device.

[0024] Fig. 3 illustrates a bottom view of an amplification device.

[0025] Figs. 4 illustrate a fluid path of an amplification device.

[0026] Figs. 5A-B illustrate an operation of a manual valve.

[0027] Fig. 6 illustrates a fluid path of an amplification device.

[0028] Fig. 7 illustrates a side perspective view of a fluid path prior to valve activation.

[0029] Fig. 8 illustrates the flow path of Fig. 7 after valve activation.

[0030] Figs. 9A-B illustrate perspective views of heating arrangements of an amplification device.

[0031] Figs. 10A-B illustrate perspective views of a printed circuit board (PCB).

[0032] FIGs. 11 A-l IB illustrates a cross-sectional view of a valve system in an amplification device having an automated valve.

[0033] FIG. 12 illustrates a cross-sectional view of a valve system in an amplification device having an automated valve.

[0034] FIG. 13 illustrates an exemplary setup for measuring valve actuation force.

[0035] FIGS. 14A-14C illustrate exemplary setups for measuring wax and / or valve actuation force.

[0036] These figures disclose embodiments of the present disclosure for illustrational purposes only. In particular, the disclosure provided by the figures and description is not meant to limit the scope of protection conferred by the present disclosure.DETAILED DESCRIPTIONDefinitions

[0037] Before the present disclosure is described in greater detail, it is to be understood that the present disclosure is not limited to particular embodiments described, and as such can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present disclosure will be limited only by the appended claims.

[0038] In general, terms used in the claims and the specification are intended to be construed as having the plain meaning understood by a person of ordinary skill in the art. Certain terms are defined below to provide additional clarity. In case of conflict between the plain meaning and the provided definitions, the provided definitions are to be used.

[0039] Where ranges and / or subranges of values are provided, the ranges and / or subranges can include the endpoints of the ranges and / or subranges. Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and lower limits of these smaller ranges can independently be included in the smaller ranges and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0040] As used herein, any reference to “one embodiment” or “an embodiment” means that a particular element, feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment. The appearances of the phrase “in one embodiment” in various places in the specification are not necessarily all referring to the same embodiment.

[0041] Additionally, certain embodiments of the disclosed devices and / or associated methods can be represented by drawings which can be included in this application.Embodiments of the devices and their specific spatial characteristics and / or abilities include those shown or substantially shown in the drawings or which are reasonably inferable from the drawings. Such characteristics include, for example, one or more (e.g., one, two, three, four, five, six, seven, eight, nine, or ten, etc.} of: symmetries about a plane (e.g., a cross-sectional plane) or axis (e.g., an axis of symmetry), edges, peripheries, surfaces, specific orientations (e.g., proximal; distal), and / or numbers (e.g., three surfaces; four surfaces), or any combinations thereof. Such spatial characteristics also include, for example, the lack (e.g. , specific absence of) one or more (e.g., one, two, three, four, five, six, seven, eight, nine, or ten, etc. of: symmetries about a plane (e.g., a cross-sectional plane) or axis (e.g., an axis of symmetry), edges, peripheries, surfaces, specific orientations (e.g., proximal), and / or numbers (e.g., three surfaces), or any combinations thereof.

[0042] In addition, it is noted that, as used herein and in the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. It is further noted that the claims can be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as “solely,” “only” and the like in connection with the recitation of claim elements, or use of a “negative” limitation.

[0043] As used herein, the terms “comprises,” “comprising,” “includes,” “including,” “has,” “having” or any other variation thereof, are intended to cover a non-exclusive inclusion. For example, a process, method, article, or assembly that comprises a list of elements is not necessarily limited to only those elements but can include other elements not expressly listed or inherent to such process, method, article, or assembly. Further, unless expressly stated to the contrary, “or” refers to an inclusive or and not to an exclusive or. For example, a condition A or B is satisfied by any one of the following: A is true (or present) and B is false (or not present), A is false (or not present) and B is true (or present), and both A and B are true (or present).

[0044] As used herein, the terms “optional,” “optionally” and “may” denote that the indicated feature may be present but can also be absent. Whenever the term “optional,” “optionally” or “may” is used, the present disclosure specifically relates to both possibilities, i.e., that the corresponding feature is present or, alternatively, that the corresponding feature is absent. For example, the expression “X is optionally substituted with Y” (or “X may be substituted with Y”) means that X is either substituted with Y or is unsubstituted. Likewise, if a component of a composition is indicated to be “optional,” the present disclosure specifically relates to both possibilities, i.e., that the corresponding component is present (contained in the composition) or that the corresponding component is absent from the composition.

[0045] The term “about” can mean within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, e.g., the limitations of the measurement system. For example, “about” can mean plus or minus 5%, 10%, or 20%, per the practice in the art. In certain embodiments, the term “about” refers to being within manufacturing tolerance levels as known by one of ordinary skill in the art (e.g., AS 1163, EN 10219, ASTM A500 or G3444 / G3466 tolerance level standards). In case of doubt, encompassed within the term “about” are numbers that are insignificantly different from the stated number.

[0046] In contrast, “distal end” refers to the end of a device or instrument that is farther away from the user or operator.

[0047] Some embodiments can be described using the expression “coupled” and “connected” along with their derivatives. For example, some embodiments can be described using the term “coupled” to indicate that two or more elements are in direct physical or electrical contact. In some embodiments, the term “coupled” is also used to describe two or more elements that are in active fluidic connection. The term “coupled,” however, can also mean that two or more elements are not in direct contact with each other, but yet still co-operate or interact with each other. The embodiments are not limited in this context unless otherwise explicitly stated.

[0048] “Operatively coupled,” “operatively connected,” and “operatively attached” as used herein means connected in a specific way that allows the disclosed devices to operate and / or methods to be carried out effectively in the manner described herein. For example, operatively coupling can include removably coupling or fixedly coupling two or more aspects (e.g., the sample preparation body and the amplification module). Operatively coupling can also include fluidically coupling two or more aspects. For example, operatively coupling a sample preparation device (sample preparation body) and nucleic acid amplification device (amplification module) can also include mateably coupling two or more components. In some embodiments, operative coupling refers to the different forms of engagement, such as (but not limited to), screwing-on, twisting-together, snapping-on, press-fitting, screwing together, sliding together, etc., of two of more aspects (e.g., the sample preparation body and the cap).

[0049] As used herein, the term “activate” or “actuate” when referring to a valve refers to the change of the valve from its closed configuration to open configuration. In some embodiments, where the valve is a sealing element, it can be actuated by displacing the sealing element to allow fluidic connection through an opening it sealed.

[0050] As used herein, the term “reagent” or “reagents” refers to any chemical, including organic compounds and inorganic compounds and combinations thereof. It can be provided in gaseous, solid, or liquid form, or any combination thereof, it can be in lyophilized form, and / or it can be a component of a solution or a suspension. In some embodiments, a reagent comprises a buffer useful in methods of detecting analytes in a sample fluid or retentate. In some embodiments, a reagent comprises an anticoagulant, a diluent, a buffer, a specific binding moiety, detectable labels, enzymes and the like. In some embodiments, a reagent can alsoinclude an extractant, such as a buffer or a chemical, to extract an analyte from a sample or a sample collection device. In some embodiments, a reagent comprises a washing buffer, a lysis buffer, a neutralization buffer, or a rehydration buffer.

[0051] As used herein, the term “biological sample” is a sample containing a quantity of organic material, e.g., one or more organic molecules, such as one or more nucleic acids e.g., DNA and / or RNA or portions thereof, which can be taken from a subject. In some embodiments, the term “biological sample” and “sample fluid” are used interchangeably. In some embodiments, the sample fluid comprises a urine sample, a saliva sample, and / or a mouthwash sample. In some embodiments, the sample fluid is formed from resuspension of a solid or semi-solid sample (e.g., fecal sample or mucosal sample) in solution. In some embodiments, the sample fluid comprises a fecal sample or mucosal sample. In some embodiments, the sample fluid is formed from resuspension any sample from a user likely to contain an analyte of interest (e.g., free nucleic acids, nucleic acids for a pathogen, a cancerous cell, or precancerous cell), as would be appreciated by one of ordinary skill in the art. In some embodiments, the cancerous and precancerous cells are identified based on expression profiles, presence or absence of certain biomarkers (e.g., cancer markers or tumor markers). In some embodiments, the sample fluid is selected from the group consisting of pus, blood sample, skin sample, etc. e.g. In some embodiments, the sample fluid is selected from the group consisting of blood, urine, semen, vaginal discharge, a vaginal swab, a nasal swab, a nasopharyngeal swab, a mid-turbinate swab, fecal sample, tears, fluid excreted at wound sites or sites of inflammation, or any other clinical material that contains nucleic acids or proteins.

[0052] As used herein, the term “nucleic acid” refers to any molecule, preferably a polymeric molecule, incorporating units of ribonucleic acid, deoxyribonucleic acid or an analog thereof. The nucleic acid can be either single-stranded or double-stranded. In some embodiments, nucleic acids refers to cell free nucleic acids (e.g., circulating cell-free DNA).

[0053] In some embodiments, the terms “disease,” “condition” or “illness” refer to a sexually transmitted disease. In some embodiments, the terms “disease,” “condition” or “illness” refer to a urinary tract infection. In some embodiments, the terms “disease,” “condition” or “illness” refer to a cancer or precancerous conditions. In some embodiments, the terms “disease,” “condition” or “illness” refer to a respiratory disease. In some embodiments, the terms“disease,” “condition” or “illness” refer to any condition caused by a pathogen (e.g., a bacterium, fungus, protist, or virus / viral agent).

[0054] As used herein, the term “biological analyte” refers to a pathogen.

[0055] A “biological sample” or “bodily fluid sample,” used interchangeably herein, can be collected from a subject. In certain embodiments, a subject is a “mammal” or a “mammalian” subject, where these terms are used broadly to describe organisms which are within the class mammalia, including the orders carnivore (e.g., dogs and cats), rodentia (e.g., mice, guinea pigs, and rats), and primates (e.g., humans, chimpanzees, and monkeys). In some embodiments, the subject is a human. The term “humans” can include human subjects of all genders and at any stage of development (e.g., fetal, neonates, infant juvenile, adolescent, and adult), where in certain embodiments the human subject is a juvenile, adolescent or adult. While the devices and methods described herein can be applied in association with a human subject, it is to be understood that the subject devices and methods can also be applied in association with other subjects, that is, on “non-human subjects.”

[0056] As used herein, the term “user” refers to a human subject.

[0057] Any terms not directly defined herein shall be understood to have the meanings commonly associated with them as understood within the art of the disclosure. Certain terms are discussed herein to provide additional guidance to the practitioner in describing the compositions, devices, methods and the like of aspects of the disclosure, and how to make or use them. It will be appreciated that the same thing can be said in more than one way. Consequently, alternative language and synonyms can be used for any one or more of the terms discussed herein. No significance is to be placed upon whether or not a term is elaborated or discussed herein. Some synonyms or substitutable methods, materials and the like are provided. Recital of one or a few synonyms or equivalents does not exclude use of other synonyms or equivalents, unless it is explicitly stated. Use of examples, including examples of terms, is for illustrative purposes only and does not limit the scope and meaning of the aspects of the disclosure herein.Nucleic Acid Amplification Device

[0058] In some embodiments, a nucleic acid amplification device may be configured to receive a liquid sample, such as from a sample preparation device. A nucleic acid amplificationdevice may amplify nucleic acids of a liquid sample through one or more incubation chambers. A nucleic acid amplification device may detect a presence of a threshold amount of targeted DNA and / or RNA via one or more detection chambers. In some embodiments, detection chambers may include lateral flow assay (LFA) strips. Implementation of the present system and devices may enable for the detection of sexually transmitted diseases, such as infections caused by bacteria (e.g., chlamydia or gonorrhea), infections caused by protists (e.g., Trichomonas vaginalis), oncological conditions (e.g., cancerous or precancerous conditions which can be determined based on the presence of cancerous or precancerous cells in the urine, saliva or mouthwash), respiratory diseases (e.g., SARS, COVID-19, flu, respiratory syncytial virus (RSV)), bacterial infections (such as urinary tract infection, urethritis, cystitis, pyelonephritis, gingivitis, and / or periodontitis), fungal infections, and / or protist infections.System Overview

[0059] Fig. 1 illustrates an embodiment of a combined sample preparation device and amplification device. System 100 may include sample preparation device 104, system housing 108, and / or amplification device 112. Sample preparation device 104 may hold or otherwise contain a liquid sample. A liquid sample may include, but is not limited to, saliva, urine, or other bodily fluids. Sample preparation device 104 may be configured to receive a liquid sample from a user and process the liquid sample, such as through a lysing process. In some embodiments, sample preparation device 104 may be configured to mix a liquid sample with one or more chemicals. For instance, sample preparation device 104 may be configured to mix a cell lysate sample to be homogeneously distributed within a buffer.

[0060] In some embodiments, sample preparation device 104 may be provided with preprocessing agents for mixing with a liquid sample. Sample preparation device 104 may provide a mixed and / or processed sample to amplification device 112. Sample preparation device 104 may mechanically and / or fluidically connect with an inlet portion of amplification device 116. In some embodiments, sample preparation device 104 may include a breakable seal that, once broken, may provide an outlet for a liquid sample that may allow a liquid sample to flow from an outlet of sample preparation device 104 to an inlet of amplification device 112. For instance and without limitation, a user may push down on a top side of sample preparation device 104, which may cause a breaking of a seal of sample preparation device 104, allowing a liquid sample to flow to an inlet of amplification device 120. Sample preparation device 104 andprocessing of a liquid sample may be as described in US Application No. 18 / 877985 and PCT Application No. PCT / EP2024 / 088263, the relevant disclosures of which are incorporated herein in its entirety.

[0061] System 100 may include amplification device 112. Amplification device 112 may be configured to receive a liquid sample from sample preparation device 104. Amplification device 112 may be designed to receive a liquid sample and detect one or more infections that can cause illness of a user. For instance, amplification device 112 may be configured to perform an amplification and detection process of one or more nucleic acids, described in further detail below. Amplification device 112 may be stored within system housing 108. System housing 108 may be made out of a plastic or other material. In some embodiments, system housing 108 may connect sample preparation device 104 with amplification device 112. For instance and without limitation, sample preparation device 104 may snap into an interior portion of system housing 108. Sample preparation device 104 may be positioned at a top of system housing 108 while amplification device 112 may be positioned at a bottom of system housing 108, which may allow one or more fluids to flow from sample preparation device 104 to amplification device 112.

[0062] In some embodiments, system 100 may include valve 120. Valve 120 may be a physical valve, as depicted in FIG. 1. In other embodiments, valve 120 may be internal to amplification device 112. For instance and without limitation, valve 120 may be an actuated valve, electromechanically operated valve, thermally activated valve such as but not limited to a wax valve, or other valve types. In some embodiments, valve 120 may be a manual valve, which may allow a user to operate valve 120 from an outside of amplification device 112. Valve 120 may be a knob or other device that may be operable to be inserted and / or rotated into an interior of amplification device 112. In some embodiments, valve 120 may be a spool type valve. Upon activation of valve 120, a fluidic communication between incubation and detection chambers of amplification device 112 may be provided, described in further detail below.

[0063] Referring still to Fig. 1, system housing 108 may include window 116. Window 116 may be positioned in front of detection chamber housing 124. In some embodiments, window 116 may have a same width and / or length of detection chamber housing 124. Window 116 may be transparent, which may allow a user to look at one or more detection chambers within detection chamber housing 124. Detection chamber housing 124 may be designed to hold one ormore lateral flow assay (LFA) strips, in some embodiments. Detection chamber housing 124 may be designed to hold two or more vertically oriented LFA strips. One or more vertically oriented LFA strips may be inserted into detection chamber housing 124 prior to an assembly of system 100. For instance, one or more vertically oriented LFA strips may be placed into detection chamber housing 124 prior to connecting system housing 108 and / or sample preparation device 104 to amplification device 112. Detection chamber housing 124 may be described in more detail below. A user may view detection chambers housed within detection chamber housing 116, which may provide an indication of a presence of a threshold amount of a nucleic acid (DNA / RNA) amplified from the sample.

[0064] Referring now to Fig. 2 A, a front perspective view of amplification device 200 is presented. Amplification device 200 may have a length of between about 30 mm to about 50 mm and / or a width of about 30 mm to about 50 mm. Amplification device 200 may have a height of about 40 mm to about 56 mm. In some embodiments, amplification device 200 may have a length of about 55 mm, a width of about 46 mm, and a height of about 56 mm. Amplification device 200 may include inlet port 204. Inlet port 204 may be circular or ovular. A shape of inlet port 204 may be designed to mate with an outlet of a sample preparation device. Inlet port 204 may include hollow opening 224. Hollow opening 224 may allow a flow of a liquid sample from a sample prep device into one or more internal components of amplification device 200. In some embodiments, hollow opening 224 may have a diameter of about 1 mm to about 2 mm. In some embodiments, inlet port 204 may include one or more sealing elements that may be adapted to seal a fluidic and / or mechanical connection between inlet port 204 and an outlet of a sample preparation device. One or more sealing elements may include an O-ring, in some embodiments. Inlet port 204 may include piercing element 216. Piercing element 216 may be conically shaped, in some embodiments. Piercing element 216 may protrude from a top surface of inlet port 204 by about 1 mm to about 4 mm. In some embodiments, piercing element 216 may have a diameter of about 3.3 mm which may taper towards a tip of piercing element 216, which may have a radius of about 0.5 mm. Piercing element 216 may be designed to pierce a seal of a sample preparation device, which may allow for a transfer of a liquid sample into inlet port 204. In some embodiments, piercing element 216 may be located at a center of inlet port 204.

[0065] As noted above amplification device 200 may include detection chambers 208. Amplification device 200 may include one or more detection chambers 208 which may receive one or more test strips. Detection chambers 208 may be made from a same mold of amplification device 200, for instance in a case of injection molding, without limitation. Detection chambers 208 may be about positioned about 40 mm to about 50 mm away from inlet port 204. In some embodiments, detection chambers 208 may include a first chamber and a second chamber. A first chamber of detection chambers 208 may be a same size of a second chamber of detection chambers 208. In some embodiments, a first chamber of detection chambers 208 may be a different size than a second chamber of detection chambers 208. Each chamber of detection chambers 208 may be formed to hold a same test strip. In some embodiments, a first chamber may be designed to hold a first type of test strip and a second chamber may be designed to hold a second type of test strip, the second type different than the first. In some embodiments, the test strip may be vertically oriented with respect to the plane of the base of the amplification device 200.

[0066] Still referring to FIG. 2A, amplification device 200 may include valve 212. Valve 212 may be a rotating, spool, or other valve. For instance, valve 212 may be adapted to be inserted into valve slot 220. Valve slot 220 may be circular, semi-circular, or other shapes. Valve slot 220 may be molded from a same mold of amplification device 200 and / or detection chambers 208, in some embodiments. Valve slot 220 may extend from inlet port 204 through detection chambers 208 away from inlet port 204. An insertion of valve 212 into valve slot 220 may cause a mixing between one or more incubation chambers and one or more detection chambers, as described in further detail below. Valve 212 may be positioned underneath test strip housing 208. For instance, valve 212 may extend along a central axis of amplification device 200 towards inlet port 204. Valve 212 and / or valve slot 220 may be positioned underneath a bottom surface of detection chambers 208.

[0067] Referring to Fig. 2B, a rear perspective view of amplification device 200 is shown. Inlet port 204 may include one or more grooves 228. Grooves 228 may be designed to hold one or more sealing elements. For instance, in some embodiments, grooves 228 may be O-ring grooves. O-rings may be placed on grooves 228, in some embodiments. Grooves 228 may be separated into two or more grooves. For instance and without limitation, a first groove ofgrooves 228 may be designed to mate with a sealing element, while a second, third, and fourth groove may be designed for retention in an injection mold.

[0068] Referring now to Fig. 3, a bottom perspective view of amplification device 300 is presented. Amplification device 300 may include all prior components described with reference to Figs. 1-2. In some embodiments, amplification device 300 may include front film 304 and / or bottom film 308. Front film 304 and / or bottom film 308 may be made out of plastic. In some embodiments, front film 304 may be welded to detection chambers 312 and bottom film 308 may be welded to an entirety of a bottom of amplification device 300. Front film 304 and / or bottom film 308 may be made out of clear acrylic, in some embodiments. Front film 304 and / or bottom film 308 may be laser welded, ultrasonic welded, heat welded, induction welded, adhesive welded, or other welding methods to amplification device 300. Front film 304 may be clear which may allow viewing of one or more vertically oriented test strips, such as vertically oriented LFA strips positioned within detection chambers 312. In some embodiments, front film 304 may act as a window, such as window 116 as described above with reference to FIG. 1. Bottom film 308 may be metallized in some embodiments, which may improve thermal conductivity of amplification device 300. For instance, a bottom of amplification device 300 may be connected to a heating arrangement and a metallization of bottom film 308 may allow for more efficient heat transfer to incubation chambers of amplification device from one or more heating elements of the heating arrangement.Fluid Pathways of an Amplification Device

[0069] Referring now to Fig. 4, a bottom view of amplification device 400 is presented. Amplification device 400 may include fluid inlet 404. Fluid inlet 404 may be in fluidic and / or mechanical connection with inlet port 204 as described above with reference to Figs. 2A-B. Fluid inlet 404 may be in fluidic communication with distribution channel 408. Distribution channel 408 may be designed to transfer a flow of fluid from fluid inlet 404 to fluid paths 424. Fluid paths 424 may provide a fluidic pathway from distribution channel 408 to incubation chambers 416. Fluid paths 424 may be about 0.8 mm to about 1 mm wide by about 0.6 to about 1 mm deep, in some embodiments. In some embodiments, amplification device 400 includes a set of incubation chambers 416, such as a set of two or more incubation chambers 416. In some embodiments, a single incubation chamber 416 may be used. Amplification device 400 may include one or more bubble traps 412. Bubble traps 412 may be designed to capture air bubblesthat may be formed within a fluid flow from fluid inlet 404 to fluid paths 424 via distribution channel 408. Amplification device 400 may have one or more detection pathways 440. Detection pathways 440 may provide a fluidic pathway between amplification chambers 416 to one or more detection chambers, such as LFA test strips.

[0070] Amplification device 400 may include valve 432, which may be as described above. Valve 432 may be inserted into a portion of amplification device 400, such as a valve chamber or inlet. In some embodiments, valve 432 may include pin 444. Pin 444 may be rectangular, ovular, triangular, or other shapes. Pin 444 may come into contact with a mechanical switch of a printed circuit board (PCB). Upon coming into contact with pin 444, a mechanical switch of a PCB board may enable a timer to of the PCB board to track an amount of time passed since an activation of valve 432.

[0071] Amplification device 400 may include one or more air bladders 420. Air bladders 420 may be designed to hold an amount of air, for instance and without limitation, between about 1 ml to about 5 ml of air. Air bladders 420 may be rigid in some embodiments. In other embodiments, air bladders 420 may be made of a flexible material that may expand and / or decompress. In some embodiments, each air bladder 420 may be separated by a wall or divider. A wall or divider may separate each air bladder 420 into symmetric shapes. For instance, each air bladder 420 may be able to expand and / or decompress over a same volume and / or may be divided to store a same volume of air. Air bladders 420 may be configured to expand while receiving air from a sample preparation device via fluid inlet 404 and / or from one or more incubation chambers. Air bladders 420 may be in fluidic communication with vents 428 via venting paths 436. Vents 428 may be hydrophobic and may prevent fluid flow via hydrophobicity. In some embodiments, vents 428 may be a sintered polytetrafluoroethylene (PTFE) frit. Vents 428 may have a thickness of between about 1 mm to about 5 mm. Vents 428 may have a diameter of about 1 mm to about 4 mm. In some embodiments, vents 428 may have a thickness of about 3 mm and a diameter of about 3.2 mm. Each vent of vents 428 may have the same dimensions. In some embodiments, each vent of vents 428 may have differing dimensions. Vents 428 may have a porosity, for instance and without limitation, of about 20 um to about 50 um. A porosity of vents 428 may be selected to increase or decrease pressure within amplification device 400. For instance, a higher porosity of vents 428 may allow for a slower pressurized loading of one or more chambers of amplification device 400 while a lower porosityof vents 428 may increase a pressurized loading of one or more chambers of amplification device 400. Each of vents 428 may have a same or differing porosity, without limitation. Air bladders 420 may provide a pressurized environment to amplification device 400, which may cause fluid to flow from fluid inlet 404 to one or more paths of amplification device 400. In some embodiments, vents 428 may allow for a transfer of air within amplification device 400 to a surrounding environment of amplification device 400 while preventing liquid from escaping amplification device 400.

[0072] A bottom of amplification device 400 may have a symmetry across a horizontal axis, in some embodiments. For instance, except for the detection pathways 440, every other element shown in a bottom of amplification device 400 may be mirrored across a horizontal axis.Amplification of a Liquid Sample

[0073] Referring still to FIG. 4, amplification device 400 may be configured to amplify a liquid sample to produce one or more amplicons. A liquid sample may be in a form of a lysate, without limitation. Each incubation chamber 416 may be design to hold an amount of a liquid sample and / or amplify the amount of the liquid sample. Amplification of a liquid sample in each incubation chamber 416 may take place using loop-mediated isothermal amplification (LAMP) or reverse transcription loop-mediated isothermal amplification (RT-LAMP) or other isothermal nucleic acid amplification method. LAMP (and RT-LAMP) may utilize specific temperatures for amplification of one or more nucleic acids. For instance, incubations chambers 416 may include one or more nucleic acid amplification compositions. Nucleic acid amplification compositions may be adapted to amplify nucleic acids of a biological analyte. In some embodiments, nucleic acid amplification compositions may be adapted to target DNA or RNA specific to one or more pathogens that can cause infections and / or illnesses. A specificity of each nucleic acid amplification composition may be based on one or more primers, such as, but not limited to, oligonucleotides, which may be specific to certain sequences of DNA and / or RNA present within a liquid sample of an subject. A primer may be configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis, or a human control sample. Nucleic acid amplification compositions may include, but are not limited to, buffers, enzymes, DNA polymerase, reverse transcriptase, salts, deoxyribonucleotides, and / or ribonucleotides. Nucleic acid amplification compositions may be provided in a form of lyophilized pellets. For instance, and withoutlimitation, nucleic acid amplification compositions may be freeze-dried and provided within incubation chambers 416. Freeze-dried pellets including one or more nucleic acid amplification compositions may dissolve upon contact with a liquid sample. In a case of a use of lyophilized pellets, lyoprotectants such as sugars and polyalchohols and / or cryoprotectants may be employed, which may preserve a function of one or more enzymes. In some embodiments, nucleic acid amplification compositions may be provided in a sphere or power.

[0074] Each incubation chamber 416 may be positioned above a heating element of amplification device 400. A heating element may be configured to increase, decrease, or maintain a temperature of each incubation chamber 416. Heating elements may include, but are not limited to, resistive, chemical, or other types of heating elements. In some embodiments, each incubation chamber 416 may have a same temperature. In other embodiments, each incubation chamber 416 may have a differing temperature. Heat generated by heating elements positioned under incubation chambers 416 may allow for an amplification of DNA and / or RNA to take place. For instance, incubation chambers 416 may be heated to temperature of about 75° C, about 70° C, about 65° C, about 55° C, preferably 50° C, or about 60° C. In some embodiments, temperatures of incubation chambers 516 may reach between about 50° C to about 80° C, between about 55° C to about 75° C, or between about 60° C to about 70° C. Incubation chambers 516 may be maintained within a target range of temperatures, which may include any range of temperatures described above, without limitation. Amplification of one or more nucleic acids may produce one or more amplicons. Amplicons may include nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

[0075] In some embodiments, amplification of nucleic acids may include nucleic acid hybridization. Nucleic acid hybridization may involve a process of annealing DNA or RNA molecules to complementary DNA or RNA molecules. In some embodiments, a doublestranded DNA sequence may be used. In other embodiments, a single-stranded DNA sequence may be used.

[0076] In some embodiments, labeling of amplified DNA and / or RNA may be performed simultaneously during amplification. For instance, nucleic acid amplification compositions may include labeling microparticles or fluorescent sequences which may be detected using fluorescence microscopy. Amplified DNA and / or RNA may be labelled through usage ofLAMP primers. In some embodiments, amplified DNA and / or RNA may be labeled with Biotin, Digoxygenin, or FAM. Oligonucleotide and / or DNA probes may be part of nucleic acid amplification compositions as described throughout this disclosure. Presence of amplified DNA and / or RNA may be assessed by measuring a turbidity of the sample in the incubation chamber before and / or after amplification.Manual Valve Embodiments

[0077] In some embodiments, the amplification device comprises a valve that is a manual valve. As used herein, the term “manual valve” refers to a valve that is manually actuated by a user. In some embodiments, the valve is partially insertable into the amplification device.

[0078] Referring now to Fig. 5A, an illustration of amplification device 500 in a closed configuration is presented. Amplification device 500 may include first incubation outlet 508 and second incubation outlet 520. First incubation outlet 508 may be positioned proximate first detection outlet 512. First detection outlet 512 may be positioned between first atmosphere outlet 516 and first incubation outlet 508. Second incubation outlet 520 may be positioned proximate second detection outlet 524. Second detection outlet 524 may be positioned between second atmosphere outlet 528 and second incubation outlet 520. Incubation outlets 508 and 516 may be each in respective fluidic communication with an incubation chamber of amplification device 500. Each detection outlet 512 and 524 may each be in fluidic communication with a detection chamber and / or a test strip such as an LFA strip, which may be vertically oriented. Each atmosphere outlet 516 and 528 may be in communication with a surrounding atmosphere of amplification device 500. For instance, in some embodiments, each atmosphere outlet 516 and 528 may include a hydrophobic vent that may allow air to escape from atmosphere outlets 516 and 528.

[0079] Still referring to Fig. 5 A, in a closed configuration, valve 504 may be partially inserted into amplification device 500, which may allow for communication of sealed volume 540 with incubation outlets 508 and 516. In some embodiments, sealed volume 540 may be in communication with detection outlets 512 and 524. Valve 504 may include a distal portion 536 that may be inserted into amplification device 500. Distal portion 536 may include a plurality of sealing elements 532. Plurality of sealing elements 532 may seal off fluid from entering one or more outlets described above.

[0080] Referring to Fig. 5B, amplification device 500 in an open configuration is presented. Valve 504 may be fully inserted into a portion of amplification device 500, which may remove sealing elements 532 from contacting outlets 508, 512, 516, 520, 524, and / or 528. A removal of one or more sealing elements 532 from one or more of outlets 508, 512, 516, 520, and / or 524 may allow for a fluidic communication between two or more of the prior listed outlets. In some embodiments, a small annular gap may be presented between a chip of amplification device 500 and valve 504, which may allow a fluid transfer between two or more outlets. For instance, in some embodiments, atmosphere outlets 528 and 516 may be sealed off while incubation outlets 508 and / or 520 may become fluidically connected to detection outlet 512 and / or 524. A connection between incubation outlets and detection outlets may allow for a transfer of a liquid sample to one or more testing strips, such as LFA strips which can be vertically oriented. A pressure difference generated by inserting valve 504 into amplification device 500 may drive fluid flow from incubation outlets 508, 520 to detection outlets 512, 524, in some embodiments. For instance and without limitation, pressure received by a sample preparation device may cause fluid to flow from incubation outlets 508, 520 to detection outlets 512, 524 as incubation outlets 508, 520 to detection outlets 512, 524 have a same air pressure as in the sample preparation device. Air pressure received by a sample preparation device may be about 0.5 bar to about 1.0 bar above an ambient atmospheric pressure. In some embodiments, fluid may flow from incubation outlet 508 to detection outlet 512 and from incubation outlet 520 to detection outlet 524. In some embodiments, fluid may flow between any incubation outlets and detection outlets described above, in any combination.Automated Valve Embodiments

[0081] In some embodiments, valve 120 may be internal to system 100. For instance, valve 120 may be thermally activated, such as a wax actuator valve (also referred to as a “wax valve”). System 100 may include a heating element which may be configured to provide an amount of heat to valve 120 in embodiments where valve 120 may be a wax valve. Heating elements may include, but are not limited to, radiative heaters, resistive heating elements, or other forms of heating elements. One or more heating elements may be positioned below and / or above valve 120. For instance in some embodiments, heating elements may be positioned below valve 120 and may apply an amount of heat to valve 120. In some embodiments, one or more heating elements may be positioned above valve 120 and may be configured to provide heat tovalve 120. Heat may be generated by one or more heating elements and may cause a wax valve to expand, thereby moving or actuating a valve in the channel. In some embodiments, this allows for fluidic communication between two or more fluid channels and / or sources described herein. Valve 120 may be a wax valve that may be positioned directly in any fluid paths and / or fluidic channels described herein, which may prevent a flow of fluid therein.

[0082] In some embodiments, heating elements of system 100 may be connected to one or more electronic components, such as but not limited to, microcontrollers, microprocessors, timers, resistors transistors, or other electronic components. A microcontroller or other processing device may activate one or more heating elements over a period of time. For instance, a microcontroller or other processing device may activate one or more heating elements after a period of about 1 minute to about 30 minutes or greater. In some embodiments, the same heating elements that may be used in LAMP cycles described herein may also be utilized for actuating the wax valve. For instance, a LAMP cycle may occur via one or more heating elements at a first temperature and after a period of time, a microcontroller or other processor may cause the one or more heating elements to increase to a second temperature greater than the first temperature, which may expand the wax valve and move it a new position within the channel.

[0083] In some embodiments, system 100 may include a space to capture melted wax in embodiments where valve 120 is a wax valve. For instance a compartment, container, or other object capable of holding liquids and / or solids may be positioned beneath valve 120 and may capture melted wax.

[0084] In some embodiments, the wax valve comprises a valve such as a ball, duckbill, umbrella, or other form of valve and an amount of wax (wax actuator). In some embodiments, the valve and wax actuator are adjacent to one another. In some embodiments, the valve is disposed within the wax actuator. As used herein, a “wax actuator” may be a form of wax capable of expanding under certain temperatures. The wax actuator is designed to expand above a temperature threshold. In some embodiments, the wax actuator can continue to expand with heating beyond its melting point. For instance and without limitation, a wax actuator may expand by about 1% to about 20% or greater in volume. A wax actuator may be positioned in a slot between two or more fluidic channels and / or below valve 120. As a wax actuator expands due to thermal energy applied to the wax actuator, the wax actuator may move (e.g., verticallymove or displace) valve 120. The movement of valve 120 may thereby allow for fluidic connection between two or more fluidic channels. In some embodiments, system 100 may include two or more wax actuators. In some embodiments, the wax actuator comprises paraffin. In some embodiments, the wax actuator comprises one or more selected from the group consisting of paraffin, polyethylene, polyolefin, and ethylene vinyl acetate (EVA).

[0085] Referring now to FIG. 11 A, a valve system 1100 A is presented. Valve system 1100A may be positioned within a portion of an amplification device 1104. Amplification device 1104 may couple with sample preparation device 104, e.g., as described above with reference to FIG. 1.

[0086] In some embodiments, amplification device 1104 may have fluidic chamber 1108. Fluidic chamber 1108 may be an incubation chamber, reaction well, or any other fluidic chamber described herein. Fluidic chamber 1108 may be fluidically couplable to fluidic channel 1132. In some embodiments, valve 1128 may be positioned between fluidic chamber 1108 and fluidic channel 1132. Valve 1128 may be an O-ring or other form of fluid stopping device. A top portion of valve 1128 may be positioned near an outlet of fluidic chamber 1108 and a bottom portion of valve 1128 may be positioned near an inlet of fluidic channel 1132. In some embodiments, valve 1128 may be positioned on top of wax actuator 1112A. Wax actuator 1112A may be positioned near a bottom of amplification device 1104 proximate printed circuit board (PCB) 1116. Wax actuator 1112A may be made out of, but is not limited to, paraffin or other forms of wax. Wax actuator 1112A may be able to expand in size in response to an increase in temperature. Wax actuator 1112A may have a melting point of about, but not limited to, about 10 degrees Celsius to about 100 degrees Celsius. In some embodiments, heating wax actuator 1112A may cause wax actuator 1112A to expand after a period of time. Periods of time may be, but are not limited to, about 1 minute to about 1 hour or greater. Heat may be applied to wax actuator 1112A above a melting point of wax actuator 1112A via second heating element 1124. As a non-limiting example, wax actuator 1112A may expand about 15% in volume after about 30 minutes of heating past a melting point of about 60 degrees Celsius.

[0087] Wax actuator 1112A may be rectangular / cuboidal, square / cubic, cylindrical, circular, ovular, or other shapes. For instance a channel in which valve 1128 and / or wax actuator 1112A may be positioned in may be rectangular in cross section. Amplification device 1104 may be positioned above PCB 1116. PCB 1116 may have first heating element 1120 and / or secondheating element 1124. First and / or second heating elements 1120,1124 may be resistive heating elements, radiative heating elements, or other forms of heating elements. First heating element 1120 may be positioned below fluidic chamber 1108. Heat from first heating element 1120 may be applied to fluidic chamber 1108, which may help facilitate reaction and / or incubation processes described herein. Second heating element 1124 may be positioned beneath wax actuator 1112 A. Heat applied to wax actuator 1112A from second heating element 1124 may cause wax actuator 1112A to expand. An expansion of wax actuator 1112A may cause valve 1128 to move upwards , away from a bottom of amplification device 1104. A movement of valve 1128 upwards away from a bottom of amplification device 1104, which may position valve 1128 in fluidic communication with an outlet of fluidic chamber 1108. In some embodiments, a movement of valve 1128 upwards away from a bottom of amplification device 1104 may cause valve 1128 to open, providing a fluidic communication between fluidic chamber 1108 and fluidic channel 1132. An expansion of wax actuator 1112A may cause valve 1128 to change configuration or shape.

[0088] In some embodiments, expansion of wax actuator 1112A may allow for fluidic chamber 1108 to become in fluidic communication with fluidic channel 1132, which may allow fluid to flow from fluidic chamber 1108 into fluidic channel 1132. Fluidic channel 1132 may be in fluidic communication with one or more detection chambers.

[0089] Referring now to FIG. 1 IB, in some embodiments of a valve system 1100B is presented. Valve system 1100B may include wax actuators 1112B and 1112C. Wax actuators 1112B and 1112C may be structurally similar to wax actuator 1112A described above with reference to FIG. 11 A. Wax actuators 1112B and 1112C may be positioned on opposite sides of valve 1128. In some embodiments, a heating element may be positioned beneath each of wax actuator 1112B and wax actuator 1112C. Heat may be applied to wax actuator 1112B and / or 1112C, which may cause wax actuator 1112B and / or 1112C to expand. Expansion of wax actuator 1112B and / or 1112C may cause compression of valve 1128. Compression of valve 1128 may enable fluidic communication between two or more outlets 1136A-F or other fluidic channels described herein. For instance, wax actuator 1112B may expand which may cause valve 1128 to move towards outlet 1136F. This may put outlets 1136A and 1136B in fluidic communication with each other. Wax actuator 1112C may expand, which may cause valve 1128 to move towards outlet 1136A. A movement of valve 1128 towards outlet 1136A may causeoutlets 1136F and 1136E to become in fluidic communication with each other. Any combination of outlets 1136A-F may become in fluidic communication with each other through movement of valve 1128, without limitation. For instance, wax actuators 1112B and 1112C may be used instead of valve 504 described above with references to Figs. 5A-B.

[0090] In some embodiments, valve 120 may be actuated through a shape memory alloy (SMA) (also referred to interchangeably as an SMA element). For instance, a SMA may be coupled (e.g., physically coupled) to a sealing element. A sealing element may be rubber, plastic, or other materials. A sealing element may have a geometry that may fit inside an interior of one or more fluidic channels which may allow the sealing element to seal off the one or more fluidic channels. In some embodiments, the SMA contracts, alters its shape, or deflects when upon exposure to heat, which may cause a tension force to be applied on a sealing element thereby pulling the sealing element out of one or more fluidic channels. System 100 may include a sealing element slot in which a sealing element may be dropped into upon a tension force applied to the sealing element via a SMA. In some embodiments, the SMA is heated to a temperature to allow it to change shape (e.g., contract, expand, deflect, etc.). In some embodiments, that temperature to change the SMA shape ranges from about 45 degrees Celsius to about 80 degrees Celsius. In some embodiments, the temperature to change the SMA shape is about 45 degrees Celsius, about 50 degrees Celsius, about 55 degrees Celsius, about 60 degrees Celsius, about 65 degrees Celsius, about 70 degrees Celsius, about 75 degrees Celsius, or about 80 degrees Celsius.

[0091] In some embodiments, a SMA may be coupled to one or more valves 120. For instance, valve 120 may be a ball, duckbill, or other form of valve that may be actuated via a contraction of a SMA. For instance and without limitation, a SMA may be exposed to heat and may contract, causing valve 120 coupled to the SMA to open or otherwise actuate, allowing fluid to flow through one or more fluidic channels. In some embodiments, a SMA may be a torsional spring inserted into a fluidic channel, thereby sealing the fluidic channel. Upon exposure to heat, a SMA may rotate out of a fluidic channel, which may unseal the fluidic channel. In some embodiments, an axial travel may be placed on top of a SMA and may move downwards with contraction of the SMA, unsealing a fluidic channel. In some embodiments, a SMA may be coupled to a flap. A flap may seal off a fluidic channel. A SMA may contract and / or rotate upon exposure to heat which may cause a flap to open, allowing fluid to flowthrough the fluidic channel. Any implementation of valve 120 described herein may be utilized with the SMA.

[0092] Referring now to FIG. 12, an embodiment of a valve system 1200 is presented. Valve system 1200 may include amplification device 1104, fluidic chamber 1108, fluidic channel 1132, PCB 1116, first heating element 1120, and second heating element 1124, each of which may be as described above with reference to FIGS. 11 A-B.

[0093] Valve system 1200 may include valve 1204. Valve 1204 may be movable within fluidic channel 1132. Fluidic channel 1132 may be directly coupled to an outlet of fluidic chamber 1108. In some embodiments, a first side of valve 1204 may be sealing fluidic chamber 1108 from fluidic communication with fluidic channel 1132 while a second side of valve 1204 may be coupled to SMA 1208. SMA 1208 may be made out of, but is not limited to, nitinol, copper-aluminum-nickel, copper- zinc-aluminum, iron- based, or other materials. SMA 1208 may be reactive to thermal energy provided by second heating element 1124. For instance, at temperatures of about 10 degrees Celsius to about 100 degrees Celsius between about 1 minute to about 30 minutes, SMA 1208 may change shape. A retraction of SMA 1208 may provide a force on valve 1204, which may open valve 1204. In some embodiments, SMA 1208 may pull valve 1204 into valve compartment 1212. Valve compartment 1212 may be an opening inside of amplification device 1104. Fluid may flow into valve compartment 1212 upon an unsealing of valve 1204 from an outlet of fluidic chamber 1108. In some embodiments, fluid may flow into valve compartment 1212 and may fill valve compartment 1212, which may cause fluid to move from valve compartment 1212 into fluidic channel 1132. Forces applied to valve 1204 via SMA 1208 may be about 1 N to about 100 N or greater. SMA 1208 may have a length of about 10 mm to about 50 mm. Valve 1204 may be a ball valve, duckbill valve, O-ring valve, or other types of valves. An opening of valve 1204 via a force caused by a change of shape of SMA 1208 may put fluidic chamber 1108 in fluidic communication with fluidic channel 1132. In some embodiments, SMA 1208 may provide a spring resistance to valve 1204 which may aid in sealing fluidic chamber 1108 from fluidic channel 1132.

[0094] In some embodiments, SMA 1208 may be implemented as a torsional spring. For instance, SMA 1208 may be formed in a spring shape and placed within a fluidic channel. Upon an application of heat, SMA 1208 in a spring shape may expand against an interior of a fluidic channel, which may cause a rotation of SMA 1208. A sealing element, such as valve 1204, maybe placed on top of an end of torsional spring version of SMA 1208, which may allow SMA 1208 to lower or lift the sealing element, sealing or unsealing fluidic chamber 1108 from fluidic communication with fluidic channel 1132. For instance and without limitation, SMA 1208 may be in a torsional spring shape and a sealing element may be placed on top of SMA 1208. A sealing element may have a key way in a first orientation that seals off fluidic chamber 1108 from fluidic communication with fluidic channel 1132 and a second orientation that provide coupling of fluidic chamber 1108 to fluidic channel 1132. Rotation of a torsional spring shape of SMA 1208 may move a sealing element from a first orientation to a second orientation and / or lift the sealing element. Alignment of a second orientation of a sealing element with fluidic chamber 1108 may allow for fluid to flow through a keyhole of the sealing element from fluidic chamber 1108 and into fluidic channel 1132. In some embodiments, valve 1204 may be a flap, such as but not limited to a silicone flap. SMA 1208 may rotate, which may cause a silicon flap version of valve 1204 to open a fluidic pathway between fluidic chamber 1108 and fluidic channel 1132. In some embodiments, two or more heaters may be placed along a length of SMA 1208 and may be configured to provide heat to different portions of SMA 1208 which may cause the different portions of SMA 1208 to contract.

[0095] Non-limiting examples of SMA include Nickel-titanium (Nitinol), gold-cadmium alloy, copper-based alloys (such as copper-zinc-aluminum and copper-aluminum-nickel), and iron-based alloys (such as iron-manganese-silicon). Other examples of shape memory alloys are further described in P. K. Kumar, et al., “Introduction to shape memory alloys ” In: Shape Memory Alloys. Springer, Boston, MA (2008).Air Bladders and Lateral Flow Assays for Nucleic Acid Detection

[0096] Referring now to Fig. 6, an example fluid path 608 within amplification device 600 is presented. Amplification device 600 may include air bladders 604. Air bladders 604 may pressurize from a filling of one or more incubation chambers 620 of amplification device 600. A pressurization of amplification device 600 may provide a force to allow fluid path 608 to form. Fluid path 608 may originate from a flow inlet of amplification device 600. Fluid path 608 may include a path towards vent 612. Vent 612 may prevent fluid flow into air bladders 604. Fluid path 608 may flow towards one or more incubation chambers 616 of amplification device 600.

[0097] In some embodiments, the present disclosure contemplates the use of one or more air bladders in a closed system — a system that lacks external venting. In some conventional device cases, external venting is preferred or instrumentation is used to assist with venting, because it is considered easier to design and / or manufacture. However, the present disclosure contemplates use of one or more air bladders, allowing for a closed system (which can be more sterile). In some embodiments, the air bladder is also larger (e.g., significantly larger than the reaction chambers. Without being bound by theory of mechanism, the larger air bladder(s) has the advantage of minimizing the backpressure in the amplification device when loading the liquid sample.

[0098] Valve 620 may be inserted, which may provide a channel for fluid pathway 608 to flow towards detection chambers 624 and / or one or more vertically oriented LFA strips.

[0099] In some embodiments, detection chambers of the present disclosure may include one or more LFA strips. In some embodiments, the LFA strips are vertically oriented relative to the base of the amplification device (e.g., on an axis perpendicular to the flow of liquid sample within the incubation chamber). LFA strips may include a sample pad, a conjugate pad, a membrane, a test line, a control line, and / or an absorbent pad and reagents for capturing and labeling amplicons, as described in more detail below. A sample pad may receive a liquid sample and may initialize a transporting of the liquid sample through the LFA strip. A liquid sample may include a lysed sample that may have one or more amplified nucleic acids (amplicons). A conjugate pad may utilize colloidal gold, colored latex beads, or fluorescent dyes. For instance, a conjugate pad may be disposed with colloidal gold particles linked to anti- FAM antibodies. In some embodiments, a conjugate pad may be disposed with labeled nucleic acid probes that may bind to a target nucleic acid present in a liquid sample.

[0100] The vertical orientation of the LFA strip(s) has several technical advantages. For example, having the LFA strips on an axis perpendicular to the flow of liquid sample within the incubation chamber allows for better control of the wicking of the amplified liquid sample from the preceding channels and incubation chamber(s). The volume of liquid in the chambers and channels is relatively high / significantly higher than that needed for the required capillary action on the LFA strips. Oversaturation of the LFA strips would be a higher risk if the LFA strips were oriented horizontally (in the same plane as the preceding channels and incubation chamber(s)). Oversaturation of LFA strips is problematic for multiple reasons (e.g., high-dosehook effect, excessive reporter concentration, excessive flow rate on the strip, inconsistent flow dynamics on the strip, oversaturation of the absorbent pad at the end of the LFA strip, backflow, etc.) and, importantly, can lead to inaccurate and / or inconclusive test results.

[0101] Alternatively or additionally, a test strip may utilize leuco dyes such as one or more of methyl green, basic fuchsin, acid fuchsin, crystal violet, malachite green or derivatives thereof. Alternatively or additionally, a test strip may utilize a rhodamine B-Cu dye which may change color in the presence of pyrophosphate. In some embodiments, one or more test strips may not be specific to any particular type of amplified nucleic acids but rather indicate an amount of DNA and / or RNA present in a liquid sample.

[0102] A membrane may be positioned between a sample pad and an absorbent pad. A membrane may have a test line and a control line. A test line may be coated with specific capture probes that may be complementary to a different region of a target nucleic acid. For instance and without limitation, Biotin labelled amplicons may be captured on a streptavidin coated line on a membrane of an LFA strip. Digoxygenin labelled amplicons may be captured on an anti-digoxygenin coated line on a membrane of an LFA strip. A target nucleic acid-probe may come into contact with a test line and may bind to one or more capture probes, which may result in a visible signal. For instance and without limitation, a target nucleic acid hybridized with biotin or digoxygenin-labelled oligonucleotide and / or FAM oligonucleotide may come into contract with a test line of an LFA strip, which may result in a visible signal indicative of a presence of a threshold amount of DNA and / or RNA. A membrane may have a control line, which may be coated with probes or antibodies that bind to one or more labeled probes or other components of a liquid sample, which may ensure that the liquid sample has migrated through the LFA strip correctly. An LFA strip may have an absorbent pad which may draw a sample through the LFA strip via capillary action and may absorb excess fluid, ensuring a continuous flow of a liquid sample. An LFA strip may be designed to indicate a presence of a threshold amount of DNA and / or RNA.

[0103] In some embodiments, systems and devices described throughout this disclosure may enable for multiplexing of LFA strips. For instance, multiple target nucleic acids may be labeled with different markers, such as colored latex beads, fluorescent dyes, gold nanoparticles, and / or other markers. A membrane of an LFA strip may have multiple test lines, each coated with various capture probes, proteins, and / or antibodies specific to a different target nucleic acid.Each test line may correspond to a specific target-probe combination. By utilizing multiplexing of LFA strips, a plurality of illness may be detected from a single liquid sample.

[0104] Referring now to FIG. 7, a side perspective view of a fluid path 700 before actuation of a valve is shown. Fluid path 700 may include fluid inlet 704, distribution channel 708, incubation chambers 712, vents 716, and / or bubble traps 720, each of which may be as described above with reference to Figs. 1-2. A sample may flow from fluid inlet 704 towards incubation chambers 712. For instance, a sample may flow from fluid inlet 704 through distribution channel 708. Distribution channel 708 may connect a single outlet of fluid inlet 704 to two or more incubation chambers 712, in some embodiments. For instance and without limitation, distribution channel 708 may provide a fluidic connection between a set of two incubation chambers 712 and fluid inlet 704. Distribution channel 708 may have two or more outlets. Each outlet of distribution channel 708 may be positioned opposite each other and / or may be mirrored across a horizontal axis. In some embodiments, each outlet of distribution channel 708 may be a same size and may allow for an equal flow of a sample from fluid inlet 704 to each incubation chamber of incubation chambers 712. In other embodiments, outlets of distribution channel 708 may be of different sizes, widths, heights, or other dimensions.

[0105] Bubble traps 720 may be positioned on each outlet of distribution channel 708. For instance, a first bubble trap 720 may be positioned on a first outlet of distribution channel 708 and a second bubble trap 720 may be positioned on a second outlet of distribution channel 708. Bubbles traps 720 may be configured to trap any bubbles that may have formed in a sample. Distribution channel 708 may connect to two or more incubation chambers 708 via fluid pathways 724. Fluid pathways 724 may include one or more fluid channels. In some embodiments, fluid pathways 724 may include a first fluid pathway and a second fluid pathway, each of which may connect to a respective incubation chamber 712. Incubation chambers 712 may connect to vents 716 via vent pathways 728. Vents 716 may be in communication with an air bladder. Vents 716 may be hydrophobic and may prevent fluid flow via hydrophobicity. Incubation chambers 712 may connect to detection pathways 736 via channel 732. For instance, each incubation chamber 712 may connect to a respective detection pathway 736 via a respective channel 732. Prior to activation by valve 744, a sample may be stored within incubation chambers 712 and may not flow in channels 732. Detection pathways 736 may flowtowards LFA contacts 740. LFA contacts 740 may be cylindrical, and may allow for a flow of a sample from incubation chambers 712 towards one or more LFA strips.

[0106] Referring now to FIG. 8, fluid path 700 as depicted in Fig. 7 after activation of a valve is presented. Activation of valve 744 may include inserting valve 744 into a valve housing. Valve 744 may include one or more sealing elements (not shown) such as, but not limited to, O-rings. For instance, valve gaps 748 may be covered by sealing elements such as O- rings. A lateral movement of valve 744 may cause channels 732 to become fluidically connected to detection pathways 736. For instance and without limitation, two or more outlets of detection pathways 736 and channels 732 may be fluidically connected between sealing elements positioned in gaps 748, as depicted above with reference to Figs 5A-B. Insertion of valve 744 may cause a pressure difference within flow path 700, which may cause a sample to flow from incubation chambers 712 to channels 732. Detection pathways 736 may be exposed to a pressure difference cause by insertion of valve 744, which may cause a sample to flow from one or more outlets of channels 732 to one or more outlets of detection pathways 736. For instance, as depicted above with reference to Figs. 5A-B, outlets of detection pathways 736 and channels 732 may be sealed together via one or more sealing elements of valve 744 positioned in gaps 748, which may expose detection pathways 736 to a pressure difference between channels 732 and detection pathways 736. A sample may flow from detection pathways 736 to LFA contacts 740 until a pressure difference within fluid path 700 has dissipated.Printed Circuit Board Components

[0107] Referring now to Fig. 9A, a top view of a PCB is presented. PCB 900 may include switch 904. Switch 904 may be configured to detect a mating or other connection to a valve of a sample preparation device, such as described above with reference to Fig. 1. Switch 904 may be a mechanical switch, in some embodiments. PCB 900 may include a timer which may be activated once a power source is connected to PCB 900, such as one or more batteries being inserted into an amplification device PCB 900 may be connected to. A processor of PCB 900 may be in communication with a timer of PCB 900 and may be programmed to determine if switch 904 is not activated within a certain time period, such as, but not limited to, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, or greater than 30 minutes. In an event switch 904 is not activated within a time period set by PCB 900, an error message may be generated by a processor of PCB 900. In some embodiments, PCB 900 may include heatingelements 908. Heating elements 908 may be resistive heating elements, in some embodiments. Heating elements 908 may include two or more heating elements 908. In some embodiments, each heating element of heating elements 908 may be positioned under an incubation chamber. Heating elements 908 may be a continuous set electrically connected via a single circuit, in some embodiments. Each heating element of heating elements 908 may be individually controlled by PCB 900. For instance, PCB 900 may adjust voltages and / or currents to each heating element 908. In some embodiments, heating elements 908 may be surface mounted on a bottom side of PCB 900, opposite a top side of PCB 900. One or more metal vias may conduct heat to a top side of PCB 900. In some embodiments, there may be a thermal conductive paste between heating elements 908 and one or more incubation chambers. PCB 900 may include one or more routed slots which may thermally isolate heating elements 904. PCB 900 may be attached to an amplification device or chip thereof via an adhesive.

[0108] Still referring to Fig. 9A, PCB 900 may include valve activation switch 912. Valve activation switch 912 may be a mechanical switch and may be activated via an insertion of a valve into an amplification device. For instance and without limitation, a portion of a valve may come into contact with valve activation switch 912, which may push a trigger of valve activation switch 912. An activation of valve activation switch 912 may trigger a timer of PCB 900. A processor of PCB 900 may be in communication with a timer of PCB 900 any may be programmed to determine if valve activation switch 912 is activated within a certain time period, such as, but not limited to, 1 minutes, 2 minutes, 3 minutes, 4 minutes, 5 minutes, or greater than 5 minutes. In an event valve activation switch 912 is not activated within a certain time period, a processor of PCB 900 may generate an error message and / or may activated light emitting diodes (LEDs) 916. LEDs 916 may be configured to emit red, blue, green, or any combination of colors thereof. In some embodiments, LEDs 916 may flash a certain color, which may indicate to a device state to a user. For instance, LEDs 916 may indicate to a user a sample preparation device was not connected to an amplification device within a certain time period, a valve was not activated within a certain time period, or other indications.

[0109] Referring now to Fig. 9B, a bottom view of a PCB 900 is shown. PCB 900 may include one or more conductors, resistors, transistors, inductors, or other electrical components. PCB 900 may include heating elements 908 as described above with reference to Fig. 9A.

[0110] Referring now to Fig. 10A, a bottom perspective view of PCB 1000 is illustrated. PCB 1000 may be PCB 900 as described above with reference to Figs. 9A-B. PCB 1000 may include heating elements 1004, which may be the same as described above with reference to Figs. 9A-B.

[0111] Referring now to Fig. 10B, a top perspective view of PCB 1000 is illustrated. PCB 1000 may include sensors 1008. Sensors 1008 may be temperature sensors, such as thermistors, in some embodiments. Sensors 1008 may be configured to generate temperature data of a respective incubation chamber, which may be communicated to a controlling element of PCB 1000. PCB 1000 may adjust heating elements 1004 based on temperature or other sensor data generated by sensors 1008.

[0112] In the following detailed examples of the present disclosure, the device will be discussed in the context of testing for sexually transmitted infections (STIs). However, it will be apparent to those skilled in the art that the device could also be employed in testing for other types of infections that can produce other illnesses, including urinary tract infections, screening for cancer, etc. Additionally, the nucleic acid amplification device as described herein may also be referred to as a cassette.

[0113] While the invention(s) has been illustrated and described in detail in the drawings and foregoing description, such illustration and description are to be considered illustrative or exemplary and non-restrictive; the invention(s) is thus not limited to the disclosed embodiments. Variations to the disclosed embodiments can be understood and effected by those skilled in the art and practicing the claimed invention(s), from a study of the drawings, the disclosure, and the appended claims.Kits

[0114] The embodiments disclosed herein also include kits including the subject devices and which can be used according to the subject methods. The subject kits can include two or more, e.g., a plurality, three or less, four or less, five or less, ten or less, or fifteen or less, or fifteen or more, of the devices or device components assay assembly components disclosed herein, according to any of the embodiments described herein, or any combinations thereof.

[0115] The kits can include one or more compositions and / or reagents, such as any of those described herein can be stored in the kits in containers separate from the devices. In addition,the kits can include any device or other element which can facilitate the operation of any aspect of the kits. For example, a kit may comprise the amplification device and instructions for use. In some embodiments, a kit may comprise the sample preparation device and instructions for use. In some embodiments, a kit may comprise any combination of the preceding.

[0116] In certain embodiments, the kits which are disclosed herein include instructions, such as instructions for using devices. The instructions for using devices are, in some aspects, recorded on a suitable recording medium. For example, the instructions can be printed on a substrate, such as paper or plastic, etc. As such, the instructions can be present in the kits as a package insert, in the labeling of the container of the kit or components thereof (i.e., associated with the packaging or subpackaging etc.). In other embodiments, the instructions are present as an electronic storage data file present on a suitable computer readable storage medium, e.g., Portable Flash drive, CD-ROM, diskette, on the cloud, etc. The instructions can be storable and / or reproducible within one or more programs, such as computer applications. The instructions can take any form, including complete instructions for how to use the devices or as a website address with which instructions posted on the world wide web can be accessed.

[0117] In some embodiments, the amplification device comprises a reusable component, e.g., substrate. In some embodiments, prior to assembly of the devices or systems provided herein, a user obtains the sample preparation device in separate packaging from the amplification device and in separate packaging from the cap. This maintains the sterility of the separate components. In some embodiments, prior to assembly of the devices or systems provided herein, a user obtains the reusable component of the amplification module separate packaging than other system components. This allows the reusable components to be sold separately from the other single-use components.

[0118] In some embodiments, a user can obtain the sample preparation device and cap in the same packaging as the amplification device.Additional Considerations

[0119] All references, issued patents and patent applications cited within the body of the specification are hereby incorporated by reference in their entirety, for all purposes.

[0120] The foregoing description of the embodiments of the disclosure has been presented for the purpose of illustration — it is not intended to be exhaustive or to limit the disclosure tothe precise forms disclosed. Persons skilled in the relevant art can appreciate that many modifications and variations are possible in light of the above disclosure.

[0121] Finally, the language used in the specification has been principally selected for readability and instructional purposes, and it may not have been selected to delineate or circumscribe the inventive subject matter. It is therefore intended that the scope of the disclosure be limited not by this detailed description, but rather by any claims that issue on an application based hereon. Accordingly, the disclosure of the embodiments of the disclosure is intended to be illustrative, but not limiting, of the scope of the disclosure.ADDITIONAL EXEMPLARY EMBODIMENTS

[0122] In one aspect, the disclosure provides a nucleic acid amplification device as contemplated in the present disclosure.

[0123] In one aspect, the disclosure provides a nucleic acid amplification device for testing a bodily fluid sample for a biological analyte, the device comprising a housing comprising: an inlet port for receiving a sample derived from a bodily fluid; an incubation and detection section configured to enable the amplification and the detection of nucleic acids, the incubation and detection section comprising: a set of incubation chambers, the set including at least two incubation chambers, wherein each incubation chambers includes a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; and one or more detection chambers; a plurality of fluid pathways connecting the inlet port with each of the incubation chambers; a plurality of detection pathways connecting each incubation chambers to the one or more detection chambers; a valve separating the set of incubation chambers and the one or more detection chambers, the valve operable to open a fluidic pathway between the set of incubation chambers and the one or more detection chambers; and a heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers, the one or more heating elements configured to provide heat to each incubation chamber of the set of incubation chambers.

[0124] In some embodiments, the valve is operable to switch the incubation and detection section between a closed and open configuration, wherein in the closed configuration fluid from the set of incubation chambers is prevented from flowing towards the one or more detection chambers and wherein in the open configuration fluid from the fluid from the set ofincubation chambers flows towards the one or more detection chambers. In some embodiments, the valve is operable to switch the incubation and detection section between a closed and open configuration via an insertion of the valve into the housing. In some embodiments, the valve comprises: a distal end insertable into the housing; and a plurality of sealing members, the plurality of sealing members positioned to seal off the fluid pathway between the set of incubation chambers and the one or more detection chambers.

[0125] In some embodiments, a printed circuit board (PCB) electrically connected to an actuator of the valve, the PCB programmed to automatically acuate the valve to open the fluidic pathway between the set of incubation chambers and the one or more detection chambers. In some embodiments, the actuator comprises a heating element and the valve comprises a wax valve meltable by the heating element.

[0126] In some embodiments, an air bladder stored within the housing, the air bladder configured to receive air from an incubation chamber of the set of incubation chambers.

[0127] In some embodiments, a hydrophobic vent in fluidic communication with the air bladder, the hydrophobic vent is configured to release an amount of air stored in the air bladder. In some embodiments, the hydrophobic vent comprises a sintered Porex Cirtek PD 103032 (PTFE) frit. In some embodiments, the hydrophobic vent has a thickness between about 1 mm to about 5 mm and a length of about 1 mm to about 5 mm. In some embodiments, the hydrophobic vent is a circular, square, or rectangular shape. In some embodiments, the hydrophobic vent has a porosity of about 20 pm to about 50 pm.

[0128] In some embodiments, a set of two air bladders, wherein each incubation chamber of the set of incubation chambers is connected to a respective first outlet in fluidic communication with a respective air bladder and a respective second outlet in fluidic communication with a respective detection chamber.

[0129] In some embodiments, the inlet port is adapted to mate with a sample preparation device. In some embodiments, the inlet port receives pressurized air from the sample preparation device, the pressurized air enabling a flow of the sample from the inlet port to the set of incubation chambers. In some embodiments, the pressurized air is about 0.5 bar to about 1.0 bar above an ambient air pressure. In some embodiments, the heating arrangement is configured to heat each of the incubation chambers to a respective target temperature.

[0130] In some embodiments, the detection chambers comprise one or more lateral flow assay (LFA) strips. In some embodiments, the LFA strips are vertically oriented with respect to a base of the housing.

[0131] In some embodiments, a bubble trap positioned at each outlet of a distribution channel connected to the inlet port and the set of incubation chambers.

[0132] In some embodiments, each fluid pathway of the fluid pathways has a cross section of between about 0.4 mm wide to about 2 mm wide by about 0.3 mm wide to about 2 mm wide.

[0133] In some embodiments, the inlet port comprises a piercing element adapted to pierce a seal of a sample preparation device. In some embodiments, the piercing element is conical. In some embodiments, the inlet port comprises a sealing element.

[0134] In some embodiments, the set of incubation chambers are positioned symmetrically across a horizontal axis of the housing.

[0135] In some embodiments, each nucleic acid amplification composition comprises a different primer configured to amplify nucleic acids indicative of a different illness. In some embodiments, a primer of a first nucleic acid amplification composition is configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a second nucleic acid amplification composition is configured to amplify nucleic acids of a another one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a third nucleic acid amplification composition is configured to amplify nucleic acids of a further one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

[0136] In some embodiments, the incubation and / or detection section comprises a leuco dye, preferably a leuco dye configured to turn to a blue color to enable detection, and / or wherein the incubation and / or detection section comprises a rhodamine B-Cu dye which changes color in the presence of pyrophosphate.

[0137] In some embodiments, the heating arrangement is configured to provide a maximum temperature of between about 50° C to about 70° C.

[0138] In some embodiments, the nucleic acid amplification compositions are provided in the form of at least one lyophilized pellet, at least one sphere, or as a powder. In some 1embodiments, the amplification device is configured to receive a sample that is a liquid suspension configured to dissolve the lyophilized pellet, sphere or powder on contact. In some embodiments, the lyophilized pellet, sphere or powder comprises a nucleic acid amplification primer configured to amplify nucleic acids indicative of at least one illness. In some embodiments, the pellet, sphere or powder further comprises an enzyme, lyoprotectant and / or a cryoprotectant.

[0139] In some embodiments, each nucleic acid amplification composition comprises a different set of primers configured to amplify nucleic acids indicative of a different pathogen.

[0140] In some embodiments, primers of the plurality of nucleic acid compositions are configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

[0141] In some embodiments, the heating element is an electrical heating element. In some embodiments, the heating element is a resistive-type heating element. In some embodiments, the amplification device further comprises at least one battery for providing energy to the heating element, preferably wherein the battery is received in the housing and / or attached to the housing. In some embodiments, the heating element is a chemical heating element, the heating element being configured to produce a chemical reaction and to heat the at least one incubation chamber to a target temperature by said chemical reaction. In some embodiments, one or more temperature sensors configured to detect a temperature of each incubation chamber of the set of incubation chambers.

[0142] In some embodiments, the bodily fluid is urine and / or saliva.

[0143] In one aspect, the disclosure provides a method of detecting a presence of a biological analyte using an amplification device, comprising: receiving a sample (e.g., bodily fluid sample) through an inlet port of the amplification device; flowing the sample to a set of incubation chambers of the amplification device from the inlet port fluid pathways, wherein each incubation chamber of the set of incubation chambers comprises a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; heating the set of incubation chambers via a heating arrangement of the amplification device, the heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers; activating a valve of the amplification device, the valve separating the incubation chamber from a detection chamber of theamplification device; flowing the sample from the incubation chamber to the detection chamber through the valve; and indicating a presence of a threshold amount of DNA and / or RNA via the one or more detection chambers.

[0144] In some embodiments, receiving the sample through the inlet port further comprises receiving pressurized air from a sample preparation device, the pressurized air having an atmospheric pressure of about 0.25 bar to about 0.75 bar above an ambient surrounding air pressure. In some embodiments, the pressurized air enables a flow of the sample to the set of incubation chambers. In some embodiments, receiving air from an incubation chamber in an air bladder; and venting excess air through a hydrophobic vent in fluidic communication with the air bladder. In some embodiments, activating the valve comprises inserting the valve into a housing of the amplification device. In some embodiments, upon insertion, the valve provides a fluidic pathway between the set of incubation chambers and the one or more detection chambers. In some embodiments, the valve is a spool valve. In some embodiments, the valve comprises: a distal end insertable into the housing; and a plurality of sealing members, the plurality of sealing members positioned to seal off the fluid pathway between the set of incubation chambers and the one or more detection chambers.

[0145] In some embodiments, activation of the valve comprises activating an actuator of the valve via a printed circuit board (PCB) electrically connected to the actuator of the valve, the PCB programmed to automatically actuate the valve to open a fluidic pathway between the set of incubation chambers and the one or more detection chambers. In some embodiments, the actuator comprises a heating element and the valve comprises a wax valve meltable by the heating element. In some embodiments, heating each of the incubation chambers to a respective target temperature.

[0146] In some embodiments, the detection chambers comprise LFA strips. In some embodiments, the LFA strips are vertically oriented.

[0147] In some embodiments, pathway of the fluid pathways has a cross section of between about 0.4 mm wide to about 2 mm wide by about 0.3 mm wide to about 2 mm wide.

[0148] In some embodiments, piercing a seal of a sample preparation device via a piercing element of the inlet port, wherein upon piercing the seal the sample flows from the sample preparation device to the inlet port. In some embodiments, the piercing element is conical.

[0149] In some embodiments, each nucleic acid amplification composition of the plurality of nucleic acid amplification compositions comprises a different primer configured to amplify nucleic acids indicative of a different pathogen. In some embodiments, a primer of a first nucleic acid amplification composition is configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, a primer of a second nucleic acid amplification composition is configured to amplify nucleic acids of a another one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a third nucleic acid amplification composition is configured to amplify nucleic acids of a further one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

[0150] In some embodiments, activating or deactivating the heating element via a mechanical switch in electrical communication with an electrical circuit between a power supply of the heating arrangement and the heating element. In some embodiments, heating comprises heating the set of incubation chambers to a maximum temperature of 75° C.

[0151] In some embodiments, the nucleic acid amplification compositions are provided in the form of at least one lyophilized pellet, at least one sphere, or as a powder.

[0152] In some embodiments, the sample is derived from a bodily fluid. In some embodiments, the bodily fluid is urine or saliva.

[0153] In one aspect, the disclosure provides a method of manufacturing and using an amplification device, comprising: injection molding a housing component out of a thermoplastic to form: an inlet port; a set of incubation chambers; and a plurality of detection chambers; fluid pathways from the inlet port to the set of incubation chambers; detection pathways from the set of incubation chambers to the one or more detection chambers; inserting a plurality of nucleic acid amplification compositions into each of the incubation chambers, the plurality of nucleic acid amplification compositions each including a primer configured to amplify nucleic acids indicative of a presence of a biological analyte; mechanically connecting a heating arrangement to a bottom portion of the housing component, the heating arrangement positioned beneath the set of incubation chambers, the heating arrangement comprising a one or more heating elements adjacent the set of incubation chambers; receiving a sample through the inlet port of the amplification device; flowing the sample to the set of incubation chambers from the inlet port through the fluid pathways; heating the set of incubation chambers via the heatingarrangement of the amplification device; activating a valve of the amplification device, the valve separating the set of incubation chambers from the one or more detection chambers; flowing the sample from the set of incubation chambers to the one or more detection chambers through the valve; and indicating a presence of a threshold amount of DNA and / or RNA via the one or more detection chambers.EXAMPLES

[0154] Below are examples of specific embodiments for carrying out the present disclosure. The examples are offered for illustrative purposes only, and are not intended to limit the scope of the present disclosure in any way. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation should, of course, be allowed for.

[0155] The practice of the present disclosure will employ, unless otherwise indicated, conventional methods of protein chemistry, biochemistry, recombinant DNA techniques and pharmacology, within the skill of the art. Such techniques are explained fully in the literature. See, e.g., T.E. Creighton, Proteins: Structures and Molecular Properties (W.H. Freeman and Company, 1993); A.L. Lehninger, Biochemistry (Worth Publishers, Inc., current addition); Sambrook, et al., Molecular Cloning: A Laboratory Manual (2nd Edition, 1989); Methods In Enzymology (S. Colowick and N. Kaplan eds., Academic Press, Inc.); Remington's Pharmaceutical Sciences, 18th Edition (Easton, Pennsylvania: Mack Publishing Company, 1990); Carey and Sundberg Advanced Organic Chemistry 3rdEd. (Plenum Press) Vols A and B(1992).Example 1: Testing Force to Activate ValvesForce to Activate Valves

[0156] An experiment is conducted to test the required actuation force to open a seal in a channel. Briefly, a valve is placed on an Instron machine to measure force — an exemplary setup is illustrated in FIG. 13. The force provides an estimate for actuation in a channel. Thereafter, seals can be selected based on required force and goals such as minimizing friction.Evaluation of Wax Actuators

[0157] An experiment is conducted to measure wax actuator activation force. Briefly, a laser-cut acrylic chamber is made with a diameter to mimic a device channel. It is filled with awax, e.g., paraffin (Sigma- Aldrich), and a rod and O-Ring seal is placed thereon, as shown in FIG. 14 A. The resulting assembly is mounted on an Instron machine and the Instron machine is set to provide constant actuation at a known force to move the rod. Heat is applied to the assembly / wax, and displacement is measured.Evaluation of SMA Springs and Tension Strips

[0158] An experiment is conducted to measure actuation force from various SMAs. For example, an nitinol spring is placed in a laser cut chamber (that mimics the device channel) attached to heating element, as illustrated in FIG. 14B. The assembly is mounted on an Instron machine. Heat is applied to the chamber and the force acting on the Instron machine is measured. After a constant force is reached, the crosshead is moved and the force applied by the spring over a range of displacements is recorded.

[0159] Alternatively, a nitinol strip is clamped on an Instron machine, adjacent to a heater, as illustrated in FIG. 14C. Nitinol strips retract when heated. Heat is applied to the strip and the tension acting on the Instron machine is measured. After a constant force is reached, the crosshead is moved and the force applied by the strip over a range of displacements is recorded.

Claims

CLAIMSWhat is claimed is:

1. An amplification device for testing a bodily fluid sample for a biological analyte, the device comprising a housing comprising: an inlet port for receiving a sample derived from a bodily fluid; an incubation and detection section configured to enable the amplification and the detection of nucleic acids, the incubation and detection section comprising: a set of incubation chambers, the set including at least two incubation chambers, wherein each incubation chambers includes a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; and one or more detection chambers; a plurality of fluid pathways connecting the inlet port with each of the incubation chambers; a plurality of detection pathways connecting each incubation chambers to the one or more detection chambers; a valve separating at least one of the set of incubation chambers and the one or more detection chambers, a wax actuator or shape memory alloy (SMA) coupled to the valve, configured to change shape from a first shape to a second shape, thereby actuating the valve from a closed to open configuration; and a heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers, the wax actuator and / or SMA, the one or more heating elements configured to provide heat to (i) each incubation chamber of the set of incubation chambers and / or (ii) change the wax actuator or SMA from a first shape to a second shape.

2. The amplification device according to claim 1, wherein the housing comprises a wax actuator.

3. The amplification device according to claim 1, wherein the housing comprises an SMA.

4. The amplification device according to claim 3, wherein the housing comprises at least two SMAs in a fluidic channel, the valve (or one or more valves) is positioned between theSMAs in the fluidic channel, and the one or more heating elements provide heat to change the at least two SMAs from a first shape to a second shape, thereby compressing and actuating the valve (or one or more valves) from open to closed configuration.

5. The amplification device according to any one of claims 1-4, wherein the change of shape is expansion, contraction, or deflection.

6. The amplification device according to claim 1, further comprising an air bladder, optionally two air bladders, stored within the housing, the air bladder configured to receive air from an incubation chamber of the set of incubation chambers.

7. The amplification device according to claim 4, further comprising an air bladder, optionally two air bladders, stored within the housing, the air bladder configured to receive air from an incubation chamber of the set of incubation chambers.

8. The amplification device according to claim 6, further comprising a hydrophobic vent in fluidic communication with the air bladder, the hydrophobic vent configured to release an amount of air stored in the air bladder.

9. The amplification device according to claim 8, wherein the hydrophobic vent comprises a sintered Porex Cirtek PD 103032 (PTFE) frit.

10. The amplification device according to claim 8, wherein the air bladder is significantly larger than the incubation chamber.

11. The amplification device according to claim 8, wherein the amplification device does not vent outside of the device.

12. The amplification device according to claim 1, further comprising a set of two air bladders, wherein each incubation chamber of the set of incubation chambers is connected to a respective first outlet in fluidic communication with a respective air bladder and a respective second outlet in fluidic communication with a respective detection chamber.

13. The amplification device according to any one of claims 1-12, wherein the set of incubation chambers are positioned symmetrically across a horizontal axis of the housing.

14. The amplification device according to claim 1, wherein the detection chambers comprise one or more lateral flow assay (LFA) strips.

15. The amplification device according to claim 14, wherein the one or more LFA strips are vertically oriented with respect to a base of the housing.

16. The amplification device according to claim 1, further comprising a bubble trap positioned at each outlet of a distribution channel connected to the inlet port and the set of incubation chambers.

17. An amplification device for testing a bodily fluid sample for a biological analyte, the device comprising a housing comprising: an inlet port for receiving a sample derived from a bodily fluid; an incubation and detection section configured to enable the amplification and the detection of nucleic acids, the incubation and detection section comprising: a set of incubation chambers, the set including at least two incubation chambers, wherein each incubation chambers includes a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; and one or more detection chambers; a plurality of fluid pathways connecting the inlet port with each of the incubation chambers; a plurality of detection pathways connecting each incubation chambers to the one or more detection chambers; a valve separating the set of incubation chambers and the one or more detection chambers, the valve operable to open a fluidic pathway between the set of incubation chambers and the one or more detection chambers; an air bladder, optionally two air bladders, configured to receive air from an incubation chamber of the set of incubation chambers; and a heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers, the one or more heating elements configured to provide heat to each incubation chamber of the set of incubation chambers.

18. The amplification device according to claim 17, wherein the air bladder is significantly larger than the incubation chamber.

19. The amplification device according to claim 17, wherein the detection chambers comprise one or more lateral flow assay (LFA) strips.

20. The amplification device according to claim 19, wherein the one or more LFA strips are vertically oriented with respect to a base of the housing.

21. The amplification device according to any one of claims 1 -20, wherein each nucleic acid amplification composition comprises a different primer configured to amplify nucleic acids indicative of a different illness.

22. The amplification device according to any one of claims 1-20, wherein a primer of a first nucleic acid amplification composition is configured to amplify nucleic acids of at least one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a second nucleic acid amplification composition is configured to amplify nucleic acids of a another one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis, and a primer of a third nucleic acid amplification composition is configured to amplify nucleic acids of a further one of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium or Trichomonas vaginalis.

23. A method of detecting a presence of a biological analyte using an amplification device, comprising: receiving a bodily fluid sample through an inlet port of the amplification device of any one of claims 1-22; flowing the sample to the set of incubation chambers of the amplification device from the inlet port fluid pathways, wherein each incubation chamber of the set of incubation chambers comprises a plurality of nucleic acid amplification compositions including a primer configured to amplify nucleic acids indicative of a presence of a pathogen; heating the set of incubation chambers via a heating arrangement of the amplification device, the heating arrangement comprising one or more heating elements positioned beneath the set of incubation chambers; actuating the valve of the amplification device; flowing the sample from the incubation chamber to the detection chamber through the valve; and indicating a presence of a threshold amount of DNA and / or RNA via the one or more detection chambers.

24. A method of manufacturing and using an amplification device according to any one of claims 1-22., comprising: injection molding a housing component out of a thermoplastic to form: an inlet port;a set of incubation chambers; and a plurality of detection chambers; fluid pathways from the inlet port to the set of incubation chambers; detection pathways from the set of incubation chambers to the one or more detection chambers; inserting a plurality of nucleic acid amplification compositions into each of the incubation chambers, the plurality of nucleic acid amplification compositions each including a primer configured to amplify nucleic acids indicative of a presence of a biological analyte; and mechanically connecting a heating arrangement to a bottom portion of the housing component, the heating arrangement positioned beneath the set of incubation chambers, and optionally valve(s), the heating arrangement comprising a one or more heating elements adjacent the set of incubation chambers and / or valve(s).

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