Patent Publication Number: US-11376588-B2

Title: In vitro diagnostic device

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
     This application claims priority to U.S. Provisional Application Ser. No. 63/051,626, titled “IN VITRO DIAGNOSTIC DEVICE,” filed on Jul. 14, 2020; U.S. Provisional Application Ser. No. 63/049,452, titled “IN VITRO DIAGNOSTIC DEVICE,” filed on Jul. 8, 2020; and U.S. Provisional Application Ser. No. 63/037,595, titled “IN VITRO DIAGNOSTIC DEVICE,” filed on Jun. 10, 2020. This application incorporates the entire contents of the foregoing applications herein by reference. 
    
    
     TECHNICAL FIELD 
     Various embodiments relate generally to in vitro diagnostic devices and methods for testing for various viruses and bacteria, such as, for example,  Streptococcus.    
     BACKGROUND 
     Every year, millions of anxious parents bring their children into a clinic, urgent care facility or emergency room presenting symptoms of pharyngitis or Group A strep. This can be a time-consuming and expensive appointment—often consuming a half-day of work or school and ultimately costing the parents $150-250 in office visit charges. And for the parents that take this step of bringing their children to a medical facility for both a rapid-test diagnosis and a longer culture-based test, 80% are sent home with negative rapid-test results, with little more than instructions to provide their ailing children with rest, fluids, over-the-counter acetaminophen and a promise of follow-up if the a more thorough culture-based test comes back positive in the following days. For many, the culture-based test does come back positive a day or two later, and the parents and their children must return to the clinic to be examined in person and for medication to be prescribed. The prescription must then be filled and picked up, often at an off-site pharmacy—adding additional time-consuming and expensive steps to the diagnosis and treatment process. 
     SUMMARY 
     In some implementations, an in vitro test device includes a plunger, a housing, one or more reagent pouches, a lateral flow test strip, and a locking member. 
     The plunger may have a plunger axis; a cylindrical plunger sidewall that is parallel to the plunger axis; a plunger base that is perpendicular to the plunger axis and open in the middle to enable communication with an interior of the plunger; a piercing member that is open in the middle to enable fluid communication with the interior and that has a smaller cross-sectional area than that bounded by the one or more plunger sidewalls; a test strip channel disposed in a sidewall in the one or more plunger sidewalls; a diaphragm member disposed in the plunger; and ridges disposed on an interior wall of the plunger. 
     The housing may have a housing axis; a cylindrical housing sidewall that is parallel to the housing axis; a housing base on a housing base end, which is perpendicular to the housing axis and closed in the middle, such that the housing base and the cylindrical housing sidewall forms a liquid-impermeable vessel; and an open end opposite the housing base that slidably receives the plunger. 
     The one or more reagent pouches may be disposed inside the housing, adjacent the housing base end, in a reagent region. The lateral flow test strip may be disposed in the test strip channel. The locking member may have a first configuration and a second configuration. In the first configuration, the locking member may prevent the piercing member from impinging into the reagent region. In the second configuration, the locking member may allow the plunger to be translated into the housing such that the piercing member impinges into the reagent region. The diaphragm member may be configured to prevent reagent from the one or more reagent pouches from leaking out of the interior when the reagent has been released from the one or more reagent pouches and when the in vitro test device is positioned vertically on its plunger base. The ridges may be configured to compress a sample collection portion of a test swab, as the test swab is passed into the interior. 
     In some implementations, an in vitro test device includes a plunger, a housing, one or more reagent pouches, a lateral flow test strip and a locking member. 
     The plunger may have a plunger axis; one or more plunger sidewalls that are parallel to the plunger axis; a plunger base that is perpendicular to the plunger axis and open in the middle to enable communication with an interior of the plunger; a piercing member that is open in the middle to enable fluid communication with the interior and that has a smaller cross-sectional area than that bounded by the one or more plunger sidewalls; and, a test strip channel disposed in a sidewall in the one or more plunger sidewalls. 
     The housing may have a housing axis; one or more housing sidewalls that are parallel to the housing axis; a housing base on a housing base end, which is perpendicular to the housing axis and closed in the middle, such that the housing base and the one or more housing sidewalls form a liquid-impermeable vessel; and an open end opposite the housing base that slidably receives the plunger. 
     The one or more reagent pouches may be disposed inside the housing, adjacent the housing base end, in a reagent region. The lateral flow test strip may be disposed in the test strip channel. The locking member may have a first configuration and a second configuration. In the first configuration, the locking member may prevent the piercing member from impinging into the reagent region. In the second configuration, the locking member may allow the plunger to be translated into the housing such that the piercing member impinges into the reagent region. 
     In some implementations, the one or more sidewalls comprise a single sidewall having a generally cylindrical form. In some implementations, the one or more sidewalls comprise four sidewalls having a generally square cross section. 
     The in vitro test device may further include a diaphragm member disposed in the plunger. The diaphragm member may be configured to prevent reagent that has been released from the one or more reagent pouches from leaking out of the interior when the in vitro test device is positioned vertically on its plunger base. The test strip channel may include an opening into the interior, such that when a test strip is disposed in the test strip channel, a portion of the test strip is adjacent the diaphragm member. 
     The plunger may further include ridges disposed on an interior wall of the plunger and spaced to compress a sample collection portion of a test swab, as the test swab is passed into the interior. 
     At least one of the plunger base and housing base may include a flat edge that prevents the in vitro test device from rolling when the in vitro test device is positioned horizontally relative to a surface and the flat edge is in contact with the surface. 
     In some implementations, each of the plunger base and the housing base include a flat edge, and the in vitro test device further includes a keying mechanism to align the plunger and housing in a fixed orientation relative to each other and to a plunger axis and a housing axis. 
     In some implementations, the in vitro test device further includes indicia on at least one of the plunger base or housing base. The indicia may provide a user with instructions regarding using the in vitro test device. The indicia may include indicia to guide manipulation of the plunger relative to the housing, or a test swab associated with the in vitro test device relative to the plunger. The indicia may include indicia to guide a user with respect to a time period during which the in vitro test device is to be positioned in a specific spatial orientation. 
     In some implementations, a method of identifying the presence of an analyte includes providing an in vitro test device and a test swab; obtaining a sample using the test swab; transitioning a locking member from a first configuration to a second configuration; advancing a plunger into a housing to pierce one or more reagent pouches in a reagent region to cause reagent therein to be released and mix; inserting the test swab with an obtained sample into the interior of the plunger; rotating the in vitro test device and disposing it on a plunger base; and determining whether the analyte is present. 
     The in vitro test device may include (a) a plunger having a plunger axis; one or more plunger sidewalls that are parallel to the plunger axis; a plunger base that is perpendicular to the plunger axis and open in the middle to enable communication with an interior of the plunger; a piercing member that is open in the middle to enable fluid communication with the interior and that has a smaller cross-sectional area than that bounded by the one or more plunger sidewalls; and, a test strip channel disposed in a sidewall in the one or more plunger sidewalls; (b) a housing having a housing axis; one or more housing sidewalls that are parallel to the housing axis; a housing base on a housing base end, which is perpendicular to the housing axis and closed in the middle, such that the housing base and the one or more housing sidewalls form a liquid-impermeable vessel; and an open end opposite the housing base that slidably receives the plunger; (c) one or more reagent pouches disposed inside the housing, adjacent the housing base end, in a reagent region; (d) a lateral flow test strip disposed in the test strip channel; (e) a locking member having a first configuration and a second configuration; wherein, in the first configuration, the locking member prevents the piercing member from impinging into the reagent region, and wherein, in the second configuration, the locking member allows the plunger to be translated into the housing such that the piercing member impinges into the reagent region. 
     The method may further include agitating at least one of the test swab or the in vitro test device. Rotating the in vitro test device may include rotating after an extraction incubation period. A mixing incubation period may separate the advancing and inserting steps. Determining may include determining based on observation of a results section of the lateral flow test strip. Determining may include determining after a testing incubation period. 
     At least one of the plunger base and the housing base may include a flat edge, and the method may further include rotating the in vitro test device such that its plunger axis and housing axis are parallel to a horizontal surface, and resting the flat edge on the horizontal surface. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1A  is perspective view of an exemplary in vitro test device. 
         FIG. 1B  is an exploded view of the exemplary in vitro test device of  FIG. 1A . 
         FIG. 2A  is a perspective view of an exemplary plunger. 
         FIG. 2B  illustrates and exemplary lateral flow test strip. 
         FIG. 2C  is a perspective view of an exemplary plunger and test strip. 
         FIG. 3  illustrates an exemplary lateral flow test strip. 
         FIGS. 4A-4G  pictorially depict the operation of the exemplary test device of  FIGS. 1A and 1B . 
         FIG. 5  is a flow diagram of an exemplary method for performing a test. 
         FIG. 6  is a cross-section of an exemplary test device. 
         FIG. 7  is a perspective view of a portion of an exemplary diaphragm. 
         FIG. 8A  is a perspective view of another exemplary in vitro test device. 
         FIG. 8B  is an exploded view of the exemplary in vitro test device of  FIG. 8A . 
         FIGS. 8C-8I  illustrate additional details of the exemplary in vitro test device of  FIG. 8A . 
         FIG. 9  graphically depicts an exemplary process of using an in vitro diagnostic device. 
     
    
    
     DETAILED DESCRIPTION 
     Described herein are various implementations of in vitro test devices and kits that can be used in an at-home setting to determine the presence of certain analytes, such as, for example a carbohydrate antigen that is unique to Group A  Streptococcus  bacteria. Such bacteria can cause strep throat, impetigo, cellulitis and other skin and soft tissue infections. By detecting its presence, or confirming its absence, appropriate treatment may be coordinated—in some cases, from home, without an expensive, time-consuming, burdensome, risk-enhancing visit to a clinic. Analytes other than those associated with Group A  Streptococcus  may also be detected with the implementations described herein. 
       FIG. 1A  is perspective view of an exemplary in vitro test device  100  and its associated test swab  103 . In some implementations, the test device  100  and test swab  103  are packaged together in a “kit,” which may be sold to consumers for purposes of determining whether certain bacteria are present (e.g., group A  Streptococcus  bacteria). 
     A user may employ the test swab  103  to take a biological sample (e.g., a pharyngeal sample), then employ the in vitro test device  100  in the manner described herein to determine—via a rapid, at-home test—whether the bacteria of interest is present, such that appropriate follow-up action can be taken (e.g., a prescription antibiotic secured). 
     As shown in  FIG. 1B , the test device  100  includes a plunger  106  and a housing  148 . As shown, the plunger  106  has a plunger axis  109  and a sidewall  112  that is parallel to the plunger axis  109 . In the implementation shown, the sidewall  112  is a generally cylindrical sidewall; in other implementations, the sidewall  112  could comprise one or more distinct sidewalls, each of which is generally perpendicular to the plunger axis. For example, in some implementations, the sidewall comprises four sides and has a generally square cross-section. 
     On one end of the plunger  106  is disposed a plunger base  115 , which, in some implementations, is perpendicular to the plunger axis  109 . The plunger base  115  is open in the middle (not visible in  FIG. 1B ) to allow communication with an interior  121  of the plunger  106 . 
     Opposite the plunger base  115 , on a piercing end  127  of the plunger  106 , is disposed a piercing member  124 . The piercing member  124  is, in some implementations, an angled or sharpened protrusion that has a smaller diameter (and smaller cross-sectional surface area) than a diameter (or cross-sectional surface area bounded by the sidewalls) of the plunger  106  itself. 
     Like the plunger base  115 , the piercing member  124  is open in the middle to enable communication with the interior  121  of the plunger. In operation, the test swab  103  may be slid into and through the interior  121 , through the open middle of the plunger base  115 , through the middle of the plunger  106  and out the middle of the piercing member  124 . 
     The test device  100  further includes a housing  148  having a housing axis  151 . The housing  148  has a housing sidewall  154  that is parallel to the housing axis  151 , and a housing base  157 , on a base end  160  of the housing, which is perpendicular to the housing axis  151 . The housing base  157  is closed in the middle, such that the housing base  157  and the sidewall  154  form a closed (e.g., liquid-impermeable) vessel  163 . 
     On the housing  148 , opposite the housing base end  160 , is an open end  164 , which is configured to slidably receive the plunger  106 . That is, the plunger  106  and housing  148  are configured such that plunger  106  can slide inside the housing  148  with a relatively tight fit (in some implementations, a liquid-tight fit), yet loose enough to permit translation of the two components during operation of the device  100 . 
     In some implementations, the housing base  157  may include a flattened edge  158  to enable the device  100  to lay flat on a horizontal surface without rolling. The plunger base  115  may also have a flattened surface  116 . In some implementations, only one of the plunger base  115  or housing base  157  has a flattened edge ( 116  or  158 , respectively); in other implementations, both plunger base  115  and housing base  157  have flatted edges  116  and  158 , and the plunger  106  and housing  148  may be keyed in some manner to maintain like orientation of the two flattened edges  116  and  158 . 
     As shown, the test device  100  further includes reagent pouch  185  and reagent pouch  186 . In some implementations, the reagent pouches  185  and  186  are disposed inside the housing  148  adjacent the base end  160 , in a reagent region  188 . The reagent pouches  185  and  186  may be constructed with sidewalls that are configured to fit into the housing  148 ; in some implementations, a top and bottom surface (e.g., surfaces that are perpendicular to the housing axis  151 ) are constructed of a foil or other thin material that is easily ruptured, e.g., by translation of the piercing member  124  into the reagent region  188 . The function of the reagent pouches is explained with reference to a lateral flow test strip  166 , and with reference to a lateral flow test strip  366  in  FIG. 3 . 
       FIG. 3  illustrates an exemplary lateral flow test strip  366 . As shown, the lateral flow test strip  366  includes a sample section  369 , which receives a sample solution; and a results section  372 , where results are displayed. 
     The process of preparing a sample for application to the lateral flow test strip  366  are briefly described. A sample may be collected in various ways, depending on the type of sample and its origin. For example, a urine sample may be collected directly on the lateral flow test strip  366  (e.g., in the case of a pregnancy test); as another example, a sample, such as a pharyngeal saliva sample, may be collected on a swab (e.g., the swab  103 ) or other sample collection device. In cases in which a swab or other sample device is used to collect a sample, the sample may be extracted into a liquid media that can then be transferred to the lateral flow test strip. 
     In some implementations, the sample is extracted by an acid, such as nitrous acid. Such an acid may serve as an oxidizing agent that breaks down cell walls of an antigen-containing target bacteria (e.g.,  Streptococcus  bacteria), to release the analyte of interest. Acids that are useful in such extraction processes may be very unstable; thus, it may be important to prepare the acids immediately prior to use. 
     One way these acids can be formed is with separate reagents that are mixed immediately prior to sample extraction. For example, in some implementations, two reagents are used—one containing a nitrite salt, such as a sodium nitrite solution, and one containing an acid, such as acetic acid. In such implementations, the combination of sodium nitrite and acetic acid produce nitrous acid. In other implementations, different reagents may be employed—such as, for example, phosphoric acid, citric acid, guanidinium thiocyanate, sodium hydroxide, etc. 
     In some implementations, dyes may be added, or the reagents may be selected, such that a color change occurs when the reagents are mixed. In such implementations, the color change can provide a user with confirmation that the reagents have been mixed and are ready to receive a sample for extraction and testing. 
     In some implementations, the separate pouches  185  and  186  ( FIG. 1B ) may contain the separate components (e.g., sodium nitrite and acetic acid) necessary to make an oxidizing agent that can extract an analyte of interest. By piercing these pouches to release and combine their contents, the extraction solution can be prepared, into which a sample-containing swab can be inserted to transfer the target analyte to the solution itself. Once the target analyte is in the solution, that solution can be transferred to the later flow test strip  366 —which is now described in more detail. 
     The exemplary lateral flow test strip  366  includes a sample pad  391 —the location at which sample liquid is applied. The sample pad  391  is absorbent and may, in some cases, include buffer salts and/or surfactants to assist a sample in flowing across the lateral flow test strip  366 . Adjacent the sample pad is a conjugate release pad  392 . In some implementations, the conjugate release pad  392  includes mobile detection particles that bind to target analytes. These detection particles may also be bound to colored or fluorescent particles—colloidal gold or latex microspheres in some implementations. Thus, in the conjugate release pad  392 , conjugates are formed between target analytes and the detection particles. 
     The lateral flow test strip  366  includes a membrane that  393  that causes the liquid sample (including the extracted analyte and any conjugate formed between extracted analyte and detection particles) to flow from the sample pad  391 , towards an absorbent pad  397 . Between the sample pad  391  and the absorbent pad  397  lies a test line  394  (or, in some cases, more than one test line) and a control line  395 . 
     The test line  394  may comprise immobilized antibodies or antigens that are configured to react with target analytes, causing the detection particles to be aggregated at the test line  394 . After enough target detection particles aggregate at the test line  394 , they may be visually apparent as a line having a contrasting color relative to the membrane  393 . If target analytes are not present in the solution passing the test line  394 , no reaction occurs, and the conjugate analyte/detection particles flow past the test line  394  without aggregating into a visual line. 
     The lateral flow test strip  366  further includes a control line  395  that confirms proper capillary flow of the test solution across the membrane  393 . In some implementations, the control line  395  appears as a visual line as soon as test solution flows past—regardless of whether a target analyte is present or not. The appearance of this line may provide some confirmation that the lateral flow test strip  366  is properly functioning. 
     As shown, the sample pad  391 , conjugate release pad  392 , membrane  393  with test strip  394  and control strip  395 , and the absorbent pad  397  are all supported by a backing card  396 —typically a substrate (e.g., thick paper, cardboard, plastic, polymer, etc.) to support and configure relative to each other the components of the later flow test strip  366 . 
     Returning to  FIG. 1A , the test device  100  further includes a removable locking clip  175  that is configured to removably clamp onto a portion of the plunger  106  to prevent the plunger  106  from sliding too far into the housing  148 . In some implementations, this locking clip  175  is in place in a first configuration, prior to the test device  100  being used, and it prevents the piercing member  124  from impinging on reagent region  188  and piercing the reagent pouches  185  and  186  in that first configuration. During operation, when it is intended for the piercing member  124  to pierce the reagent pouches  185  and  186 , the removable locking clip  175  can be removed and the plunger depressed (a second configuration—in which the plunger  106  is more deeply disposed in the housing  148 , wherein the piercing member  124  is disposed in the reagent region  188 ). 
     As shown, the test device  100  also includes a diaphragm member  191  disposed in the plunger. As will be explained in more detail with reference to other figures, in some implementations, the diaphragm  191  member is configured to provide a liquid seal around a test swab  103 , when said test swab  103  is inserted into and through the plunger  106 . 
       FIG. 2A  illustrates additional details of an exemplary plunger  206 . Disposed on an exterior surface  236  of the plunger  206  (e.g., recessed in the thickness of sidewall  212 ) is a test strip channel  239  and a fluid channel  242 . The test strip channel  239  and the fluid channel  242  intersect as a sample region  245 . In some implementations, there is more than one fluid channel  242 . For example, as shown (only partially visible), a second fluid channel  242 A extends just beyond the sample region  245  and extends parallel to the fluid channel  242 , towards a piercing member  224 . 
       FIG. 2B  illustrates an exemplary lateral flow test strip  266 . The test strip channel  239  is configured to receive the lateral flow test strip  266 , which, in some implementations, includes a sample section  269 , which is configured to receive a liquid sample; and a results section  272 . 
     In some implementations, as shown in  FIG. 2C , the plunger  206  is configured such that the lateral flow test strip  266  fits into the test strip channel  239  in a manner that aligns its sample section  269  with the sample region  245  that is at the intersection of the test strip channel  239  and the fluid channel  242 . As will be described in more detail, sample fluid may flow down the fluid channel  242  and reach the lateral flow test strip  266  at its sample section  269 , at the sample region  245 . 
       FIGS. 4A-4H  pictorially depict the operation of the exemplary test device  100 . As depicted in  FIG. 4A , the test device  100  and swab can be removed from the packaging in which they came. In its packaging, the test device  100  is protected by the removable locking clip  175 . That is, the test device  100  is in a first configuration, such that the plunger  106  is maintained in the housing  148  in a manner that prevents the piercing member  124  from impinging upon the reagent region  188  and piercing either reagent pouch  186  or reagent pouch  185 . The test device may be positioned vertically, such that it is supported by the housing base  157 . 
     As depicted in  FIG. 4B , the removable locking clip  175  is removed. This facilitates additional translation of the plunger  106  within the housing  148 . 
     As depicted in  FIG. 4C , the plunger  106  is depressed, or translated more deeply into the housing  148 . The piercing member (not visible in  FIG. 4C ) impinges into the reagent region  188 , and that piercing member pierces the reagent pouch  186  and reagent pouch  185 . In some implementations, the reagent pouch  186  and reagent pouch  185  are cylindrical in shape, to match the shape of the housing  148 , and they may have rigid sidewalls; their top and bottom surfaces (those that are perpendicular to the axis  151  of the housing), however, may comprise an easily pierceable foil or other membrane. Thus, when the piercing member impinges on the reagent region  188 , both reagent pouch  186  and reagent pouch  185  may be pierced, releasing their contents into a common space and allowing those contents to mix. 
     In some implementations, the mixing of the reagents contained in the reagent pouch  186  and the reagent pouch  185  causes a solution to be formed that is suitable for extracting a sample from the swab  103 . For example, in some implementations, reagent pouch  186  contains sodium nitrite solution, reagent pouch  185  contains acetic acid, and when the two reagent pouches  186  and  185  are pierced and their contents are combined, nitrous acid is a formed—an acid that may be effective in extracting a sample from the swab  103 . 
     As depicted in  FIGS. 4D and 4E , the swab  103  (with a sample contained thereon, such as a pharyngeal saliva sample) is inserted into the interior  121  of the plunger. Once fully inserted, a tip of the swab  103  containing the sample comes into contact with the mixed contents of reagent pouch  186  and reagent pouch  185  (e.g., an “extraction solution”). This extraction solution (e.g., nitrous acid in some implementations) may cause relevant portions of the sample to be extracted from the swab  103 , into the extraction solution itself. 
     In some implementations, the housing  148  and plunger  106  are agitated for a short period of time, prior to the swab  103  being inserted. In some implementations, a short “mixing incubation period” may also be provided prior to the swab being inserted  103  (e.g., to enable the multiple reagents to fully mix). The desire or need for agitation or incubation may depend on the specific reagents employed and the nature of a target sample. 
     As depicted in  FIG. 4F , the test device  100  is rotated 180 degrees, such that it is vertically oriented and resting on its plunger base. In some implementations, this rotation occurs after an “extraction incubation period”—a period of time during which the extraction solution chemically and biologically acts on the sample to extract a target analyte. The extraction incubation period may be 30 seconds, 1 minute, 2 minutes, 3 minutes, 5 minutes, or some other period of time that facilitates extraction of sufficient quantity of analyte for use in subsequent steps. 
     As shown in one implementation, the swab  103  remains disposed in the test device  100  once it is rotated. In such implementations, the diaphragm  191  (shown in  FIG. 1B ) may prevent the extraction solution from leaking out through the interior  121  portion of the plunger  106 . Once the test device  100  is rotated, (with reference to  FIGS. 2A and 2C ) the extraction solution may be directed to the sample region  245  via the fluid channel  242 , causing the extraction solution to come into contact with the sample section  269  of the lateral flow test strip  266 . In other implementations, the test swab  103  may be removed before the test device  100  is rotated. 
     As depicted in  FIG. 4G , the test device  100  may be rotated again and placed horizontally, such that it rests, in a rotation-free manner, on a flat portion  158  of the housing base  157  and/or on a flat portion  116  of the plunger base  115 . In this position, it may be possible for a user to easily read results of the later flow test strip  266  (e.g., through a transparent or semi-transparent portion of the housing  148 ). 
     In some implementations, the test device  100  is not rotated horizontally until after a “testing incubation period”—a period of time during which (with reference to  FIG. 3 ) the extraction solution wicks from the sample pad  391 , through the conjugate release pad  392 , across the membrane  393 , and through the test line  394  and control strip  395  in the results section  372 . The testing incubation period may be 30 seconds, 1 minute, 2 minutes, 3 minutes, 5 minutes, or some other period of time that facilitates movement of the extraction solution across the lateral flow test strip  266  or  366 . 
       FIG. 5  is a flow diagram of an exemplary method  500  for performing a test using an exemplary in vitro test device. As depicted, the method  500  includes providing ( 501 ) an in vitro test device, such as, for example, the in vitro test device  100  and corresponding test swab  103 . 
     The method  500  further includes obtaining ( 504 ) a sample using the test swab. For example, a user may obtain a pharyngeal saliva sample from a patient using the test swab  103 . 
     The method  500  further includes releasing ( 507 ) the locking mechanism. For example, with reference to  FIG. 1B  and  FIG. 4B , the locking clip  175  may be removed, to enable the plunger  106  to translate relative to the housing  148 . In some implementations, the locking mechanism may have a different form than the locking clip  175  (e.g., see locking mechanism  875 , illustrated in and described with reference to  FIG. 8C ). 
     The method  500  further includes disposing ( 508 ) the device on its housing base (e.g., vertically oriented, on the housing base  157 —for example, as shown in  FIG. 4B . 
     The method  500  further includes advancing ( 510 ) the plunger into the housing. For example, with reference to  FIG. 1B  and  FIG. 4C , the plunger  106  may be advanced into the housing  148 , such that the piercing member  124  impinges on the reagent region  188 , causing the reagent pouch  186  and the reagent pouch  185  to be pierced, such that their contents mix, forming an extraction solution. In some implementations, the method  500  includes waiting ( 511 ) a “mixing incubation period” of time, to allow the reagents to fully mix with each other, to form an extraction solution. 
     The method  500  further includes inserting ( 513 ) the test swab into the plunger. For example, with reference to  FIG. 4D  and  FIG. 4E , the test swab  103  may be inserted into the plunger  106 , specifically into the interior space  121 , bringing the sample end of the test swab  103  into contact with the extraction fluid, which, as shown, will be adjacent the housing base  157 . 
     The method  500  further includes, in some implementations, agitating ( 516 ) the test device. This step could include, for example, “swirling” the test device  100  or test swab  103  in a circular motion to agitate the extraction fluid and increase its interaction with the test swab—to promote or expedite extraction of a sample contained on the test swab. The method  500  further includes waiting ( 517 ) for an “extraction incubation period” of time, to allow for sufficient sample to be extracted by and into the extraction solution, from the test swab. 
     The method  500  further includes rotating ( 519 ) the test device. For example, with reference to  FIG. 4F , the test device  100  is rotated ( 519 ) such that it is vertically oriented on its plunger base  115 . In some implementations, in this configuration, the extraction fluid—which will now include any target analyte from the test swab  103 —will flow to the lateral flow test strip (e.g., via the fluid channel  242  shown in  FIG. 2A  and  FIG. 2C , to the sample region  245  and the sample  269  on the lateral flow test strip  266 ). 
     The method  500  further includes waiting ( 520 ) for a “testing incubation period” of time. In some implementations, this period of time allows the extraction fluid to wick up the lateral flow test strip (e.g., the membrane  393 ), such that any target analyte in the extraction fluid will cause a test line  394  to appear; regardless of the presence of any target analyte, in some implementations, wicking of the extraction fluid past the control line  395  will cause a visible line to appear there. 
     The method  500  further includes determining ( 522 ) whether an analyte is present. In some implementations, this includes determining ( 522 ) whether a test line is present on the lateral flow test strip. For example, with reference to  FIG. 3 , this could include determining ( 522 ) whether two lines are present (e.g., both a test line  394  and a control line  395 )—as opposed to no lines (e.g., in the case of a faulty lateral flow test strip) or only one line (e.g., a control line  395 ). 
     In some implementations, steps in the method  500  may be reordered or omitted, or other steps may be added. For example, in some implementations, it may not be necessary to agitate ( 516 ) the test device. In some implementations, it may not be necessary to wait ( 511 ) for a mixing incubation period of time. In some implementations, the locking clip may be removed ( 507 ), the test device disposed ( 508 ) on its base, and the plunger advanced ( 510 ) prior to the sample being obtained ( 504 ); in this manner, the sample may be obtained while the user is waiting ( 511 ) for any necessary incubation period of time. In some implementations, the test swab may be removed from the test device prior to the test device being rotated ( 519 ). In such implementations, removal of the test swab facilitate a “squeezing” of the tip of the test swab by a diaphragm (e.g., the diaphragm  191  shown in  FIG. 1B ), which may cause addition sample to be extracted from the test swab. In such implementations, an additional agitation step may be added prior to the test device being rotated ( 519 ). The waiting ( 520 ) step may be omitted or inherent in the process; that is, the user may simply determine ( 522 ) whether two lines appear on the lateral flow test strip, without “waiting” ( 520 ) an explicit period of time. Other variations are possible. 
       FIG. 6  is a cross-section of the test device  100 , with the piercing member  124  of the plunger disposed in a second configuration, in which it impinges on the reagent region  188 , piercing the reagent pouch  186  and reagent pouch  185 . As shown, the test swab  103  is disposed in the interior space  121  of the plunger  106 .  FIG. 6  further illustrates how the diaphragm  191  can, in some implementations, provide a seal around the test swab  103 . In such implementations, the reader will appreciate how that seal may prevent or limit extraction solution from leaking through the interior space  121  when the test device  100  is rotated. In some implementations, the diaphragm  191  may also provide a seal that substantially prevents reagent (or much reagent) from leaking out of the interior space  121  (whether or not the test swab  103  is in place) when the test device is rotated such that its plunger base  115  is on the bottom. 
     In some implementations, such as the one shown, the diaphragm member  191  has a protrusion  692  that interfaces with an opening  693  in the plunger body (see also  FIG. 4E )—for example, to anchor the diaphragm member  191  in place in the plunger. In other implementations, the diaphragm member  191  is anchored in another manner (e.g., with adhesive, or with a multi-step or co-molding process). In still other implementations, the diaphragm member  191  is omitted. 
       FIG. 7  is a perspective view of a portion of an exemplary diaphragm member  191 . As shown, the diaphragm member includes slits  701 A,  701 B and  701 C. In some implementations, these slits  701 A,  701 B and  701 C converge at an axis  704  of the diaphragm member  191 . The diaphragm  191  may comprise a resilient material that permits passage of a test swab therethrough. The material may be both stiff enough and flexible enough to substantially seal against the test swab. In addition, the material may be stiff enough to effectively “squeeze” the test swab as it is removed from the test device  100  (e.g., to extract additional extraction fluid, with, in some cases, target analyte—to enhance the extraction process). 
       FIG. 8A  is perspective view of another exemplary in vitro test device  800  and its associated test swab  803 . In some implementations, the test device  800  and test swab  803  are packaged together in a “kit,” which may be sold to consumers for purposes of determining whether certain bacteria are present (e.g., group A  Streptococcus  bacteria). Specifically, a user may employ the test swab  803  to take a biological sample (e.g., a pharyngeal sample), then employ the in vitro test device  800  as described herein, to determine—via a rapid, at-home test—whether bacteria of interest are present, such that appropriate follow-up action can be taken (e.g., a prescription antibiotic secured). 
     As shown in  FIG. 8B , the test device  800  includes a plunger  806  and a housing  848 . As shown, the plunger  806  has a plunger axis  809  and a sidewall  812  that is parallel to the plunger axis  809 . In the implementation shown, the sidewall  812  is a generally cylindrical sidewall; in other implementations, the sidewall  812  could comprise one or more distinct sidewalls, each of which is generally perpendicular to the plunger axis. For example, in some implementations, the sidewall comprises four sides and has a generally square cross-section. 
     On one end of the plunger  806  is disposed a plunger base  815 , which, in some implementations, is perpendicular to the plunger axis  809 . The plunger base  815  is open in the middle to allow communication with an interior  821  of the plunger  806 . In some implementations, as shown, the plunger  806  is made up of two different portions—one portion including the primary sidewall  812 , and one portion including the base  815 ; such implementations may be configured to retain a diaphragm member  891 , which may be included and configured to provide a seal around a test swab  803 , when said test swab  803  is inserted into and through the plunger  806 . In other implementations, the plunger  806  may be a single unitary piece that includes both the primary sidewall  812  and the plunger base  815 . 
     Opposite the plunger base  815 , on a piercing end  827  of the plunger  806 , is disposed a piercing member  824 . The piercing member  824  is, in some implementations, an angled or sharpened protrusion (or multiple protrusions, as shown in one implementation) that has a smaller diameter (and smaller cross-sectional surface area) than a diameter (or cross-sectional surface area bounded by the sidewalls) of the plunger  806  itself. 
     Like the plunger base  815 , the piercing member  824  is open in the middle to enable communication with the interior  821  of the plunger. In operation, the test swab  803  may be slid into and through the interior  821 , through the open middle of the plunger base  815 , through the middle of the plunger  806  and out the middle of the piercing member  824 . 
     The test device  800  further includes a housing  848  having a housing axis  851 . The housing  848  has a housing sidewall  854  that is parallel to the housing axis  851 , and a housing base  857 , on a base end  860  of the housing, which is perpendicular to the housing axis  851 . The housing base  857  is closed in the middle, such that the housing base  857  and the sidewall  854  form a closed (e.g., liquid-impermeable) vessel  863 . 
     On the housing  848 , opposite the housing base end  860 , is an open end  864 , which is configured to slidably receive the plunger  806 . That is, the plunger  806  and housing  848  are configured such that plunger  806  can slide inside the housing  848  with a relatively tight fit (in some implementations, a liquid-tight fit), yet loose enough to permit translation of the two components during operation of the device  800 . 
     In some implementations, the housing base  857  may include a flattened edge  858  to enable the device  800  to lay flat on a horizontal surface without rolling. The plunger base  815  may also have a flattened surface  816 . In some implementations, only one of the plunger base  815  or housing base  857  has a flattened edge ( 816  or  858 , respectively); in other implementations, both plunger base  815  and housing base  857  have flatted edges  816  and  858 , and the plunger  806  and housing  848  may be keyed in some manner to maintain like orientation of the two flattened edges  816  and  858  (e.g., by a raised nub  881  and corresponding open slot  882 , as shown in one implementation). 
     As shown, the test device  800  further includes reagent pouch  885  and reagent pouch  886 . In some implementations, the reagent pouches  885  and  886  are disposed inside the housing  848  adjacent the base end  860 , in a reagent region  888 . The reagent pouches  885  and  886  may be constructed with sidewalls that are configured to fit into the housing  848 ; in some implementations, a top and bottom surface (e.g., surfaces that are perpendicular to the housing axis  151 ) are constructed of a foil, plastic, polymer, or other thin membrane that is easily ruptured, e.g., by translation of the piercing member  824  into the reagent region  888 , but that seals and retains reagents until the piercing member  824  releases them. 
       FIG. 8C  illustrates an exemplary locking member  875  that, in some implementations, prevents the plunger  806  from being inadvertently advanced into the corresponding housing  848 . In some implementations, the locking member  875  has two configurations—a first configuration, in which the locking member  875  extends beyond an inner diameter of the housing member  848 , thereby preventing the plunger  806  from being advanced further into the housing member  848 ; and a second configuration, in which the locking member  875  can be depressed inward, such that it does not extend beyond an inner diameter of the housing member  848 , thereby allowing the plunger  806  to be advanced deeper into the housing  848  (such that its piercing member  824 , shown in  FIG. 8B , can be translated into the reagent region  888 , for example, to pierce reagent pouches  885  and  886  disposed therein). 
     In some implementations, as shown in  FIG. 8D , the locking member  875  is a tab that results from a slot  877  being formed in the plunger base  815 , with a section  878  of material of the plunger base  815  remaining intact to form a “hinge.” A thickness and elasticity of the section  878  may influence how much force the locking  875  exerts to prevent the plunger  806  from being disposed into the housing  848 , when the locking member  875  is in a first configuration. 
     In some implementations, as shown, indicia  879  (e.g., lock symbol and/or a directional arrow) may be formed on the locking member  875 , to guide users in the operation of the device  800 . In some implementations, the locking member  875  may be colored differently, relative to the plunger  806  and/or plunger base  815 , to make the locking member  875  stand out visually (e.g., the locking member may be colored red or green, relative to a gray or white color of the plunger  806 ). 
     Other indicia may be employed on the device  800 , as shown in  FIGS. 8E, 8F and 8A . For example, guidance indicia  880  for where a swab is to be inserted in the plunger base  815  may be applied to the plunger base  815  (e.g., through molding, engraving or printing processes). Additional guidance indicia  883  may be provided on the housing base  857 , for example to remind a user to retain the device  800  in a particular position for a period of time. A company logo/name  884  may be applied to a portion of the device (see  FIG. 8A ). Other indicia may be applied. The indicia may be molded or engraved into components of the device  800  itself; or indicia may be printed or applied to, for example, a sticker or coating that is applied to a portion of the device (e.g., to provide additional guidance, user instructions, warnings, or information). 
       FIG. 8G  illustrates additional details of an exemplary plunger  806 . Disposed on an exterior surface of the plunger  806  (e.g., recessed in the thickness of sidewall  812 ) is a test strip channel  839 . In the implementation shown, the test strip channel  839  includes an opening  842  into the interior  821  of the plunger. A test strip  844  can be disposed in the test strip channel  839 , such that a sample section  843  of the test strip  844  is positioned in the interior  821  of the plunger  806 . 
     With reference to  FIG. 8H  and the preceding figures, when the test strip  844  is disposed as just described, and when the reagents are released from their pouches  885  and  886  (e.g., by impingement of the plunger  806 , particularly the piercing member  824 , into the reagent region  888 ), and when the device swap is inserted into the plunger  806  (swab not shown in  FIG. 8H ), and the device  800  flipped upside down, such that the plunger base  815  is on the bottom, the mixed reagents  845  collect against the diaphragm member  891 , such that the mixed reagents  845  are in contact with the sample region  843  of the test strip  844 . In this manner, in the implementation shown, a patient sample collected on a swab and extracted by the reagents  845  can be absorbed by the test strip  844 , and a result (e.g., a positive or negative indication of the presence of a particular analyte, such as Group A  Streptococcus  bacteria) can be obtained. 
     In some implementations, the diaphragm  891  may also provide a seal that substantially prevents reagent (or much reagent) from leaking out of the interior space  821  (whether or not the test swab  803  is in place), when the test device is rotated such that its plunger base  815  is on the bottom. 
     Turning to  FIG. 8I , some implementations of a device  800  (e.g., the plunger  806 ) include features that enhance the extraction of a sample from a test swab. In particular, in the implementation shown, ridges  850  are disposed on an interior wall of the plunger  806 . In some implementations, such ridges are configured to form an inner “diameter”  853  that is smaller than an uncompressed diameter of a tip of a corresponding swab  803 , such that as the swab  803  is inserted into the device  800  and/or removed from the device  800 , its sample-collection tip is compressed in a way that agitates or squeezes additional sample material into the reagent. 
       FIG. 9  graphically depicts a process of using an in vitro diagnostic device, such as the device  800  illustrated in and described with reference to the preceding figures. In a first step, a locking member is released, such that the plunger of the device can be translated into the housing of the device. For example, with reference to  FIGS. 8B and 8C , a locking member  875  can be released (e.g., depressed), to enable the plunger  806  to be depressed, or translated, into the housing  848 . 
     The action of depressing the plunger  806  into the housing  848  is depicted in a second step in  FIG. 9 . This step can cause a piercing member  824  of the plunger to impinge into a reagent region  888 , causing the piercing member  824  to pierce reagent pouches  885  and  886  and release the reagents therein, enabling them to mix. In some implementations, the mixing of the reagents may bring about a color change, to provide feedback to a user that the mixing has occurred and that the reagent mixture is ready to receive a biological sample. 
     In a third step depicted in  FIG. 9 , confirmation may be made that the plunger is sufficiently translated into the housing. In some implementations, indicia, such as arrows on the plunger and housing, may be closely aligned when the plunger and housing are appropriately positioned relative to each other. 
     In a fourth step depicted in  FIG. 9 , a swab may be prepared for sample collection. In some implementations, as shown, this can involve removing the swab from protective packaging in which it is provided. 
     In a fifth step depicted in  FIG. 9 , a patient from whom a sample is to be collected (in this example, a pharyngeal sample) can be directed to open his or her mouth. Optionally, a tongue depressor may be employed to hold the patient&#39;s tongue down and expose the back of the patient&#39;s throat (tongue depressor shown in second frame of a sixth step). 
     In a sixth step (first frame) depicted in  FIG. 9 , a tip of the sample collection swab can be rubbed across the back of the patient&#39;s throat and against the tonsils and tonsil area. This sixth step is illustrated from the front (first frame) and from the side (second frame). The swab can then be removed from the patient&#39;s throat (e.g., without contact with the patient&#39;s cheeks, tongue, or teeth). 
     In a seventh step depicted in  FIG. 9 , the swab can be inserted into the in vitro diagnostic device. For example, the swab  803 , after it has been used to collect a pharyngeal sample, can be inserted into the device  800 —specifically, into the interior space  821  of the plunger, through the middle opening of the plunger base  815 . The swab can be advanced all the way into the device, past the ridges  850 , to the bottom of the housing  848 , where the mixed reagent  845  sits. 
     As depicted in an eighth step in  FIG. 9 , it can be advisable, in some implementations, to agitate the swab in the reagent mixture for a period of time (e.g., 5 seconds, 10 seconds, etc.), then to leave the device undisturbed for another period of time (e.g., 30 seconds, one minute, two minutes, five minutes, etc.), such that the reagent mixture has an opportunity to extract (e.g., via a chemical reaction and mechanical dispersion) the biological sample from the tip of the swab. 
     As depicted in a ninth step in  FIG. 9 , the swab can, in some implementations, be removed from the device. In some implementations, the step of removing the swab from the device can cause additional sample material to be “squeezed” out of the swab (e.g., by the ridges  850  and/or the diaphragm member  891 ). 
     As depicted in a tenth step in  FIG. 9 , the device can be rotated 180 degrees. For example, the device  800  can be flipped from resting on its housing base  857  to its plunger base  815 . In some implementations, as illustrated in  FIG. 8H , this can cause mixed reagent  845  to move from the bottom of the housing  848 , in the reagent region  888 , to an area adjacent the diaphragm member  891  and, more importantly, to an area where the sample section  843  of the test strip  844  is positioned. In this position, the mixed reagent  845 —with its biological sample contained therein—is drawn into the test strip  844 , where presence of a particular analyte (e.g., Group A  Streptococcus  bacteria) can be detected. 
     As depicted in a eleventh step in  FIG. 9 , the device can be laid on its side. For example, the device  800  may be laid on its side, such that that the flat region  816  of the plunger base  815  and/or flat region  858  of the housing base  857  are resting on a surface, such as a table. In this position, in some implementations, the test strip  844  is visible to a user—that is, the test strip  844  is facing up or out for easy viewing. 
     In some implementations, a period of time is required for the reagent/sample mixture to be drawn into the test strip. This waiting period (e.g., one minute, two minutes, five minutes, ten minutes, etc.—as specified in instructions associated with the device) is depicted in a twelfth step in  FIG. 9 . After this period, or within aa particular time window (e.g., between five and ten minutes), the test strip can be read. For example, the test strep  366  shown in  FIG. 3  may be read to confirm the presence of a control line  395  and the presence or absence of a test line  394 . As described with reference to  FIG. 3 , the presence of the test line  394  can indicate a positive result (e.g., affirmative detection of an analyte of interest), and the absence of the test line  394  can indicate a negative result (e.g., indication that the analyte of interest was not detected). 
     While various implementations have been described with reference to exemplary aspects, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof without departing from the contemplated scope. For example, a cylindrical housing is described, but housing could take another shape, such as one with a square cross-section. Two reagents are described, but in some implementations, there may be a single reagent; in other implementations, there may be three or more reagents. A lateral flow test strip may have multiple test strips and thus be able to detect multiple analytes. One or more incubation periods may be unnecessary in some implementations; in other implementations, one or more incubation periods may be longer or shorter than specified. Agitation may be required in some implementations but not in other implementations. Analytes other than those associated with Group A  Streptococcus  may be detected. For example, some in vitro diagnostic devices may be employed to detect urinary tract infections, yeast infections, sexually transmitted diseases, other infectious diseases, etc. 
     In general, many modifications may be made to adapt a particular situation or material to the teachings provided herein without departing from the essential scope thereof. Therefore, it is intended that the scope not be limited to the particular aspects or embodiments disclosed but include all aspects falling within the scope of the appended claims.