Patent Publication Number: US-2007122912-A1

Title: Sample Substrate for Use in Biological Testing and Method for Filling a Sample Substrate

Description:
CROSS REFERENCE TO RELATED APPLICATION  
      This application is a divisional of U.S. patent application Ser. No. 10/309,311 filed Dec. 4, 2002, which is incorporated herein by reference. 
    
    
     FIELD  
      The present teachings relate to devices for storing samples to be tested. More particularly, the present teachings relate to various sample substrates for use in biological testing devices, and methods for filling a sample substrate.  
     BACKGROUND  
      Biological testing has become an important tool in detecting and monitoring diseases. In the biological testing field, thermal cycling is used to amplify nucleic acids by, for example, performing polymerase chain reaction (PCR) and other reactions. PCR may be carried out using “consumables”, which are sample substrates that are relatively inexpensive, disposable, readily available, and often having multiple sample wells, for example, such as PCR tubes, chips, sample plates, trays, or microcards, thus, enabling varying volumes of samples to be processed and tested. As mentioned above, one such consumable that may enable a number of reactions in a relatively small amount of space is commonly known as the microcard, a spatial variant of the micro-titer plate, which may contain individual wells with a wide range of volumes.  
      Microcards may be “pre-loaded” with a dried reagent or other similar element of a sample to be tested in each of the sample wells. This pre-loading may be done by the microcard manufacturer who provides the pre-loaded card to the testing facility to be further loaded with a desired test sample. Such a pre-loaded microcard may limit the capabilities of a testing facility to configure their card for a desired test to the configuration of cards they have already ordered from the manufacturer. In addition, the testing facility may be required to wait for a newly configured card to be delivered by the manufacturer, possibly delaying desired testing. Microcards in use today may be filled at the testing facility using filling devices that may be costly for smaller testing facilities to maintain. There exists a need for a low-cost consumable that may be fully configured with varying test samples by a user to a desired configuration for testing.  
     SUMMARY  
      In accordance with the teachings, a sample substrate for use in biological testing is provided having a first member defining at least one sample well and a second member including means for substantially sealing the at least one sample well. The means for substantially sealing may be movable with respect to a remainder of the second member.  
      As used herein, the term “substantially seal” refers to a state whereby a sample well is essentially closed off so that material contained within the sample well remains within the sample well, and material outside of the sample well is substantially inhibited from flowing into the sample well. “Substantially sealed” is not intended to define a state whereby no material can get in or out of the sample well, but just a state of sealing sufficient to allow a level of isolation of a sample within the sample well for desired testing to occur. By way of example only, this state of being “substantially sealed” is intended to describe a state similar to that achieved by staking, a method of sealing sample wells within a microcard by deforming a metal backing of a microcard to sufficiently isolate the sample to allow testing to occur.  
      According to another aspect, a sample substrate for use in biological testing may comprise a first member defining a plurality of sample wells for containing a sample to be tested and a second member including a plurality of sample well closure elements. Each sample well closure element may be movable with respect to a remainder of the second member. The second member may be movable with respect to the first member from an uncovered position, wherein the plurality of sample wells is uncovered, to a covered position, wherein the plurality of sample wells is substantially covered by the second member. At least one of the plurality of sample well closure elements may be configured to substantially seal a corresponding sample well when the second plate is in the covered position, by moving the at least one of the plurality of closure elements from a first predetermined position to a second predetermined position.  
      According to yet another aspect, at least one of the plurality of closure elements may comprise a cap and an annular rim surrounding the cap.  
      In another aspect, the cap may include a cylindrical portion configured to engage an inner surface of its corresponding sample well.  
      In a further aspect, the annular rim may comprise a snap-action hinge that moves the cap from the first predetermined position to the second predetermined position upon a sufficient force being imparted on the cap.  
      In yet another aspect, the annular rim may be configured to allow the at least one of the plurality of caps to move with respect to the remainder of the second member from the first predetermined position to the second predetermined position.  
      According to another aspect, a portion of the at least one of the plurality of closure elements may reside within the corresponding sample well when the closure element is in the second predetermined position.  
      In another aspect, at least one of the at least one of the plurality of closure elements and its corresponding sample well may comprise a flexible portion configured to deflect to maintain substantially the same volume of the sample to be tested within the sample well when the at least one of the plurality of closure elements is in the second predetermined position as compared to a volume of the sample to be tested when the closure element is in the first predetermined position.  
      In yet another aspect, the sample substrate may include at least one reservoir in fluid communication with the at least one of the plurality of sample wells.  
      According to another aspect, the reservoir may be in fluid communication with the at least one of the plurality of sample wells via a fluid channel.  
      In further aspect, the sample substrate may comprise a branch fluid channel between the fluid channel and the at least one of the plurality of sample wells.  
      According to yet another aspect, the at least one of the plurality of closure elements may permit fluid communication between its corresponding sample well and the reservoir when in the first predetermined position and prevents fluid communication between the reservoir and the sample well when in the second predetermined position.  
      According to another aspect, at least one reservoir may be capable of being filled with the sample to be tested when the second member is in the covered position.  
      In another aspect, the at least one reservoir may comprise a plurality of reservoirs.  
      In yet another aspect, each of the plurality of reservoirs may be in fluid communication with a separate portion of the plurality of sample wells.  
      In another aspect, at least a portion of the at least one closure element may comprise a light pipe.  
      In another aspect, a light pipe may be located on the flexible portion of the at least one closure element.  
      According to another aspect, the plurality of sample wells may be positioned in a matrix.  
      According to yet another aspect, the plurality of closure elements may be positioned in a matrix and each of the plurality of closure elements may be configured to mate with a corresponding one of the plurality of sample wells.  
      In another aspect, the sample substrate may comprise at least one of 4, 8, 12, 16, 24, 48, 96, 128, 384, and 1536 sample wells and corresponding closure elements.  
      In yet another aspect, an adhesive membrane may be positioned between the first and second member when the microcard is in the covered position.  
      According to another aspect, the adhesive membrane, before a first use of the microcard, may affixed to the first member or the second member.  
      According to yet another aspect, the first member may comprise a first plate and the second member may comprise a second plate.  
      In another aspect, the sample substrate comprises a microcard. In yet another aspect, the sample substrate comprises a micro-titer plate.  
      In another aspect, a method of filling a sample substrate may comprise placing a first material into at least one of a plurality of sample wells defined by a first member of the sample substrate, placing a second material into at least one of the plurality of sample wells, moving a second member of the sample substrate with respect to the first member to substantially cover the plurality of sample wells, and moving at least one of a plurality of closure elements comprised by the second member from a first predetermined position to a second predetermined position to substantially seal the at least one of the plurality of sample wells.  
      In yet another aspect, the first material may comprise a reagent and the second material may comprise a biological sample to be tested.  
      According to another aspect, the first material and the second material may be placed into the at least one of the plurality of sample wells before the second plate is moved to substantially cover the plurality of sample wells.  
      According to a further aspect, the first and second materials may be placed into the at least one of the plurality of sample wells via a pipette.  
      According to yet another aspect, the first material may be placed into the at least one sample well before the second member is moved to substantially cover the plurality of sample wells.  
      In another aspect, the first material may be placed into the at least one sample well via a pipette.  
      In yet another aspect, the second material may be placed into the at least one of the plurality of sample wells after the second member is moved to substantially cover the plurality of sample wells.  
      According to another aspect, the second material may be placed in a reservoir of the sample substrate and transferred to the plurality of sample wells by at least one of vacuum loading and centrifugal loading.  
      According to yet another aspect, the moving at least one of the plurality of closure elements comprises using a fixture to apply pressure to the at least one of the plurality of closure elements thus moving the at least one of the plurality of closure elements with respect to its corresponding sample well and with respect to the second member.  
      In another aspect, a portion of at least one of the plurality of closure elements may deform when the plurality of closure elements move to substantially seal its corresponding sample well.  
      In yet another aspect, a portion of at least one of the plurality of sample wells may deform when its corresponding closure element moves to substantially seal the sample wells.  
      According to another aspect, a portion of at least one of the closure elements may be at least partially submerged in the first and second materials contained in its corresponding sample well when the at least one closure element substantially seals its corresponding sample well.  
      According to yet another aspect, the submerged portion of the closure element may comprise a light pipe.  
      In one aspect, a sample substrate for use in biological testing may comprise a first member defining a plurality of sample wells, and a second member including a plurality of corresponding sample well closure elements, each of the plurality of closure elements corresponding to one of the plurality of sample wells. The second member may be movable with respect to the first member from an open position, wherein the plurality of sample wells are open, to a covered position, wherein the plurality of sample wells are substantially covered by the second member. The plurality of sample well closure elements may each be movable with respect to a remainder of the second member from a first predetermined position to a second predetermined position configured to substantially seal a corresponding sample well when the second member is in the closed position.  
      In another aspect, a sample substrate for use in biological testing may comprise a first member defining a plurality of sample wells for containing a sample to be tested and a second member including a plurality of sample well closure elements. Each sample well closure element may be movable with respect to a remainder of the second member. The second member may be movable with respect to the first member from an uncovered position, wherein the plurality of sample wells are uncovered, to a covered position, wherein the plurality of sample wells is substantially covered by the second member. At least one of the plurality of sample well closure elements may be configured to substantially seal a corresponding sample well when the second plate is in the covered position, by moving the at least one of the plurality of closure elements from a first predetermined position to a second predetermined position. The microcard, before a first use, may have sample wells containing no material to be tested and may be in the uncovered position.  
      According to another aspect, a sample substrate for use in biological testing may comprise a first member defining a plurality of sample wells for containing a sample to be tested and a second member including a plurality of sample well closure elements. Each sample well closure element may be movable with respect to a remainder of the second member. The second member may be movable with respect to the first member from an uncovered position, wherein the plurality of sample wells is uncovered, to a covered position, wherein the plurality of sample wells is substantially covered by the second member. At least one of the plurality of sample well closure elements may be configured to substantially seal a corresponding sample well when the second plate is in the covered position, by moving the at least one of the plurality of closure elements from a first predetermined position to a second predetermined position. The microcard may be in the covered position and may have material to be tested contained within at least one of the sample wells, the at least one of the plurality of sample wells being substantially sealed by the closure element.  
      In yet another aspect, a sample substrate for use in biological testing may comprise a first member defining a plurality of sample wells for containing sample to be tested and a second member including a plurality of sample well closure elements and a surface connecting the sample well closure elements. Each sample well closure element may include a cap with a projecting member dimensioned to fit into a corresponding sample well and a flexible annular hinge member connecting the cap and the surface of the second member. The flexible annular hinge member may be configured to snap between a first discrete position in which the cap substantially covers the corresponding sample well and a second discrete position in which the cap substantially seals the corresponding sample well.  
      In still another aspect, a sample substrate for use in biological testing may comprise a first plate-like member defining an array of sample wells for containing sample to be tested and a second plate-like member including an array of sample well closure elements and a surface connecting the sample well closure elements. The sample well closure elements may be positioned to correspond with the array of sample wells. Each sample well closure element may including a cap with a cylindrical member dimensioned to fit into a corresponding sample well and a bottom portion, and a flexible annular hinge member connecting the cap and the surface of the second plate-like member. The flexible annular hinge member may include an over-center hinge so that the hinge member snaps between a first discrete position in which the cap is spaced from the sample well, and a second discrete position in which the bottom portion of the cap is positioned within the sample well to substantially seal the corresponding sample well.  
      It is to be understood that both the foregoing general description and the following description of various embodiments are exemplary and explanatory only and are not restrictive. 
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
      The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate at least one exemplary embodiment. In the drawings,  
       FIG. 1  is a plan view of a microcard having  384  samples wells in an open position;  
       FIG. 2  is a plan view of the microcard of  FIG. 1  in a closed position;  
       FIGS. 3A-3D  are partial section views of a sample well of the microcard of  FIG. 1  showing a progression of steps to fill and substantially seal the sample wells;  
       FIG. 4  is a plan view of an alternative embodiment of a microcard having  96  sample wells;  
       FIG. 5  is a plan view of an alternative embodiment of a microcard in an open position;  
       FIGS. 6A-6C  are partial section views of a sample well of the microcard of  FIG. 5  showing a progression of steps to fill and substantially seal the sample wells;  
       FIG. 7  is a plan view of an alternative embodiment of a microcard; and  
       FIG. 8  is a partial section view of a sample well of an alternative embodiment of a microcard having a light pipe. 
    
    
     DESCRIPTION OF VARIOUS EMBODIMENTS  
      Reference will now be made to various exemplary embodiments, examples of which are illustrated in the accompanying drawings. Wherever possible, the same reference numbers are used in the drawings and the description to refer to the same or like parts, and the same reference numbers with alphabetical suffixes or numerical prefixes are used to refer to similar parts.  
      In accordance with various embodiments, a sample substrate is provided. In one aspect, the sample substrate may be filled with one or more samples to be tested in a testing device. Such a testing device may include a thermal cycler or other suitable biological testing device. In various aspects, the sample substrate may comprise a plurality of sample wells located in a first member, with each of the sample wells having an associated closure element located in a second member. In some embodiments, the two members may be formed of a single piece and movable with respect to one another to allow open access to the sample wells in a first (“uncovered”) position and to cover the sample wells in a second (“covered”) position.  
      It should be understood that although the term “microcard” is used in the specification, the present teachings are suitable in any type of sample substrate such as, for example, micro-titer plates, sample trays, etc. In various embodiments, such as shown in  FIGS. 1-3 , a sample substrate such as microcard  10  is provided. Microcard  10  may be configured for thermally cycling samples of biological material in a thermal cycling device. The thermal cycling device may be configured to perform nucleic acid amplification on samples of biological material. One method of performing nucleic acid amplification of biological samples is PCR. Various PCR methods are known in the art, as described in, for example, U.S. Pat. Nos. 5,928,907 and 6,015,674 to Woudenberg et al., the complete disclosures of which are hereby incorporated by reference for any purpose. Other methods of nucleic acid amplification include, for example, ligase chain reaction, oligonucleotide ligations assay, and hybridization assay. These and other methods are described in greater detail in U.S. Pat. Nos. 5,928,907 and 6,015,674, which are also incorporated herein by reference.  
      In certain embodiments, the microcard may be used with a real-time detection system. Real-time detection systems are known in the art, as also described in greater detail in, for example, U.S. Pat. Nos. 5,928,907 and 6,015,674 to Woudenberg et al., incorporated herein above. During real-time detection, various characteristics of the samples are detected during the thermal cycling. Real-time detection permits accurate and efficient detection and monitoring of the samples during the nucleic acid amplification. Alternatively, the microcard may be used in a thermal cycling device that performs endpoint detection of the nucleic acid of the samples. Additional examples of thermal cyclers used in PCR reactions include those described in U.S. Pat. Nos. 5,038,852 and 5,333,675, the contents of both of which are hereby incorporated by reference herein.  
      In  FIG. 1 , a plan view of a microcard  10  is shown in an open position and having a first member, or plate,  12  and a second member, or plate,  14 . First plate  12  and second plate  14  are connected via a hinge element  16 , which may be of the living hinge type, for example. Microcard  10  may be made formed as a single unit out of a material such as polypropylene that is both suitable for PCR testing and for comprising a living hinge. Other materials may also be used that are capable of providing the proper characteristics suitable for use in a PCR testing device. Although in certain embodiments it may be desirable for microcard  1   0  to be formed as a single piece it may also be possible to form plates  12  and  14  as separate pieces joined by a hinge element that may be integral with one of plates  12  or  14  and attached to the other or a separate element attached to both.  
      As shown in  FIG. 1 , plate  12  defines a plurality of sample wells (or sample chambers)  20   a - 20   c.  As embodied herein, plate  12  defines 384 sample wells divided into three sets of 128 sample wells. As shown in  FIG. 1 , each set of 128 wells is configured in a 8×16 matrix. It should be understood that a wide variety of configurations are possible. Other common configurations include, for example, 48, 96, and 384 sample well matrices, although the present teachings are suitable with any number of sample wells. Plate  12  also defines a plurality of channels  22   a - 22   c  that connect sample wells  20   a - 20   c  via branch channels  26   a - 26   c  so as to be in fluid communication with a respective reservoir  24   a - 24   c.  Although three reservoirs  24   a - 24   c  are depicted, each in fluid communication with one-third of the  384  sample wells, other configurations are possible. For example, one reservoir may be provided that is in fluid communication with all of the sample wells  20  or there may be twenty-four reservoirs, each in communication with one of the channels. Further, any other number of reservoirs may be contemplated so as to be in communication with a desired portion of sample wells.  
      A sample substrate such as a microcard may be “spotted” with a reagent in one or more of the sample wells, which is then dried. As used herein, spotting defines the process of placing a fluid, for example a reagent, into a sample well, often using a pipette, but other suitable filling means may be employed. These pre-loaded microcards may then be filled with another fluid, for example a biological sample to be tested, so as to create a reaction between the reagent and the sample during the PCR process. Similar to microcard  10 , traditional microcards may have one or more reservoirs that may be filled with the sample to be tested. The sample fluid contained in the reservoirs may then pass to the sample wells, for example, by vacuum loading or by centrifugal loading, whereby the card is spun in a centrifuge to transfer the liquid from the reservoir to the sample wells with which the reservoir communicates, as well as any other means known in the art for loading the sample wells with a biological sample.  
      Microcard  10  may be used in a somewhat similar fashion, but because it allows a user open access to each individual well, it may provide more flexibility in how the microcard is configured. For example, a user may spot a reagent of his or her choice into one or more of the sample wells  20   a - 20   c  when the microcard is in the open or “uncovered” position shown in  FIG. 1 . Microcard  10  may be configured so as to be compatible with automatic pipetting equipment or it may be suited for manual pipetting or other spotting means. Such a user configurable card may allow the user to decide at the time of testing what samples and reagents to use in the testing rather than relying on pre-loaded cards.  
      The user may also introduce a variety of reagents into the sample wells. As depicted in  FIG. 1 , for example, a user may introduce  128  separate reagents into each of sample wells  20   a  when the microcard is in the uncovered position. Reservoir  24   a  could then be filled with a biological sample that could react with each of the different reagents during PCR testing. This procedure could be repeated for loading reagents into sample wells  20   b,    20   c  and a separate biological sample could be placed in each of reservoirs  24   b  and  24   c.  Such a configuration could then be used, for example, to screen three individuals for a variety of diseases or other conditions. In addition, by spotting each of wells  20   a - 20   c  with different reagents, a single biological sample could be loaded into each of reservoirs  24   a - 24   c.  Thus, with the microcard depicted in  FIGS. 1-3 , a single sample could be screened for 384 different properties.  
      In another testing configuration, each well could be loaded with a separate biological sample and one or more of the reservoirs could be loaded with a single reagent or separate reagents. This configuration, which may be referred to as a reverse card, could allow for screening of a single condition in a variety of biological samples. For example, a population could be screened for the existence of one condition. The various configurations of loading microcards described herein are merely exemplary. Other configurations both of reservoir number and sample/reagent loading in the sample wells may be apparent from the teachings of the disclosure contained herein.  
      Once the reservoirs  24   a - 24   c  have been filled and the sample wells  20   a - 20   c  have been appropriately spotted, plate  14  may then be folded over onto plate  12  as seen in  FIG. 2 , which is shown at an outside surface of plate  12 .  FIG. 2  shows a covered position wherein the plurality of sample wells  20   a - 20   c  are substantially covered by plate  14 . Although reservoirs  24   a - 24   c  are depicted as being fully covered by plate  14 , it is possible in certain embodiments for reservoirs  24   a - 24   c  to be provided with an opening (not shown) so that reservoirs  24   a - 24   c  may be filled after plate  14  is moved into position over plate  12 . The opening may be in the form of a through hole located in either of plate  12  or plate  14  so as to allow access by a pipette or other filling means, or as is possible with centrifugally loaded microcards, the reservoir may be open substantially along an edge at the periphery of the card.  
      With traditional microcards, the sample wells are often provided in a polypropylene card, although other PCR compatible materials besides polypropylene may be used. A foil backing may be adhered to the card to close off each of the sample wells, channels, and/or reservoirs thus maintaining the desired separation between various of the reservoirs, sample wells, and reservoirs. In order to provide a similar isolated covering, an adhesive membrane  30  (see  FIGS. 3A-3D ) may be provided between plates  12  and  14 . Adhesive membrane  30  may be made of a material such as polypropylene, LEXAN, MYLAR, or any other suitable PCR-compatible material. Adhesive membrane  30  may be initially affixed to either of plates  12  or  14  with an adhesive backing to provide the desired seal between plates  12  and  14  once microcard  10  is in the closed position. As depicted in  FIGS. 3A-3D , membrane  30  is initially affixed to plate  14  and moves into contact to adhere with plate  12 . Membrane  30  is preferably configured to adhere to plate  12  so that fluid communication between reservoirs  24   a - 24   c  and sample wells  20   a - 20   c  via channels  22   a - 22   c  and branch channels  26   a - 26   c  is maintained when plates  12  and  14  are adhered together. Membrane  30  may be coated with a PCR-compatible adhesive, such as one that is non-fluorescing and has high-tack properties. It is desirable that membrane  30  be configured so as not to inhibit fluid flow from reservoirs  24   a - 24   c  to each of the sample wells  20   a - 20   c.    
      Plate  14 , which may be moved into position over plate  12 , comprises a plurality of closure elements  40 , as shown, for example, in  FIG. 3B . Each closure element  40  is configured to be positioned relative to a corresponding sample well  20  so as to substantially cover and then substantially seal the sample well once it has been filled with the desired fluids for reaction during PCR testing. In various embodiments, the closure element  40  comprises a flexible annular rim  42  and a cap  44 . In the embodiment shown in  FIG. 3B , the flexible annular rim  42  defines a hinge that connects plate  14  to cap  44 . The flexible annular rim  42  surrounds cap  44 , but permits axial movement of cap  44  during a closing procedure described below.  
      In various embodiments, cap  44  comprises a cylindrical member  45  and a bottom member  47 . The cylindrical member  45  extends downward from a top surface  48  of the cap  44 . The cylindrical member  45  includes an outer surface  49  preferably dimensioned to closely fit or have an interference fit with the inner cylindrical surface of the sample well  20  to substantially seal the sample well when the cap is moved downward into the sample well. The cap  44  may move downward by an external force being placed on the top surface  48  of the cap, causing the annular rim (or hinge)  42  to pivot so that the cap  44  moves axially in the sample well  20 . The annular rim  42  shown in  FIGS. 3B-3D  is configured so that it snaps downward from a first predetermined (or discrete) position ( FIGS. 3B-3C ) to a second predetermined (or discrete) position ( FIG. 3D ) downward from the first predetermined position. The annular rim  42  may define an over-center hinge that will maintain the cap in either of two predetermined (or discrete) positions: a first position ( FIGS. 3B-3C ) or a second position ( FIG. 3D ).  
       FIGS. 3A-3D  show a sequential operation of spotting, closing, filling, and substantially sealing one of sample wells of microcard  10  of  FIG. 1  (for simplicity, the a-c designation has been dropped in reference to elements  20 ,  22 ,  24 , and  26  in  FIGS. 3A-3D ). As embodied in  FIG. 3A , sample well  20  located in plate  12  has been spotted with a reagent  50 . This may be done prior to or after placing the plate  14  on plate  12 . As seen in  FIG. 3B , plate  14  may then be moved into a position (also called the “covered” position) over plate  12  by, for example, rotating plate  14  about hinge element  16  and pressing on plate  14 . Adhesive membrane  30  may provide a seal between plates  12  and  14 , but may maintain an open fluid path via channel branch  26 , which connects to channel  22  and ultimately to reservoir  24 .  FIG. 3B  shows the closure element  40  and cap  44  in a first position. In various embodiments, the first position is a discrete predetermined position of the hinge (or annular rim)  42 .  
       FIG. 3C  shows the closure element  40  and cap  44  still in the first position. As shown in  FIG. 3C , fluid  60  contained in reservoir  24   a  has flown into sample well  20  via channel branch  26  due to, for example, a vacuum or centrifugal force, thereby mixing with reagent  50  in sample well  20 . Once the desired fluids and reactants have been combined in sample well  20 , cap  44  may be moved into a second position within sample well  20  to substantially seal, or isolate, sample well  20  from channel branch  26 , as shown in  FIG. 3D . In the example shown, the cap  44  may be moved to a second position by a user pressing downward on the top surface  48  of cap  44  with a sufficient force to cause the bottom portion of the cap to slide axially into sample well  20 . Alternately, any type of pressing mechanism may be used to push downward on the top surface  48  of cap  44 .  
      The hinge (or annular rim)  42  is configured so that the closure element  40  (including cap  44 ) snaps from the first position (shown in  FIGS. 3B-3C ) to the second position (shown in  FIG. 3D ) upon the lowering of the cap beyond a certain predetermined point. Once the cap  44  is in the second position, the cap is sufficiently lowered so that bottom member  47  of cap  44  blocks the channel branch  26 , therefore preventing fluid communication between the channel branch  26  and the sample well  20 . The outer surface  49  of the cylindrical member  45  of cap  44  may be configured to have a close clearance with an inner surface of the sample well  20 . The engagement of the outer surface  49  of cap  44  with the inner surface of the sample well promotes substantial sealing between cap  44  and sample well  20 . Caps  44 , for example, could be moved into the substantially sealed position individually or substantially all at once.  
      In certain embodiments, the bottom member  47  of the cap may be provided with a flexible portion. As shown in  FIG. 3D , the bottom member  47  may include a flexible portion  46 . Likewise, as also shown in  FIG. 3D , the portion of plate  12  defining sample well  20  may also be provided with a flexible portion  20 - 1 . Flexible portions  46  and  20 - 1  compensate for the fluid, a combination of reagent  50  and sample  60 , contained within sample well  20  as cap  44  is moved into position to substantially seal sample well  20  by bulging in opposite directions to maintain substantially the same fluid volume within sample well  20 . As used herein, “substantially the same volume” is intended to refer to the volume of material contained in the sample well before and after cap  44  is moved into place to substantially seal sample well  20 . Substantially the same volume is not intended to mean that the volume within the sample well remains exactly the same, and is intended to allow for some amount of material to possibly flow out of sample well  20  as cap  44  is moved into place. By incorporating flexible portions  46  and  20 - 1  into microcard  10 , cap  44  and sample well  20  are capable of compensating for at least some of the sample material that would otherwise be displaced by cap  44  as it moves into place within the sample well. With a microcard of the present teachings, radiation may be directed to a detecting device either through cap  44  or through the bottom of sample well  20  depending on the configuration of the PCR testing device used.  
      During PCR testing, undesirable condensation may form within the sample well and obscure a viewing window into sample well  20  through which radiation, e.g., fluorescence, may pass and be detected by the PCR testing apparatus. An advantage achieved by various embodiments of a microcard according to the present teachings is that cap  44  may be inserted within sample well  20  so that a portion, for example flexible portion  46 , is in contact with the sample. With a portion of cap  44  in direct contact with the sample, radiation may more easily pass through plate  14  without being affected by any potential condensation within sample well  20 .  
      In addition, with conventional devices, it may be necessary to stake the sample wells after they have been filled with the desired reactants. In the case of a microcard with a foil backing, this is often accomplished by deforming a metal backing with a stylus or other suitable device so that the foil backing protrudes into a feed channel, such as channel branch  26 , and blocks it so that it is no longer in fluid communication with its feed channel and reservoir. Closure element  40  may perform this function of substantially sealing sample well  20  through its snap-fit into well  20 , thus eliminating the need to stake the microcard.  
      In order to move caps  44  into the substantially sealed position, a fixture may be provided that could contact the top surface  45  of caps  44  and press the caps into position within sample wells  20 . This same fixture could be provided as a two-stage press that is also capable of aligning and mating plates  12  and  14  before microcard  10  is filled via a centrifugal or vacuum fill, for example. Plates  12  and  14  may fit together via an interference fit whereby one of plates  12  and  14  has a rim configured to fit around a periphery of the other of plates  12  and  14  with the interference fit being sufficient to hold the two plates together. Other snap-fit means such as snap tabs as well as any other suitable closure means may be employed to fit plates  12  and  14  together. It also may be desirable to heat one plate and cool the other plate to achieve a temporary size difference between the two plates  12 ,  14 . Plates  12  and  14  may then be moved into a closed position and, as their temperatures equalize, a tight interference fit may be achieved. The fixture used may be configured to provide this selective temperature difference between the two plates.  
      As is clear from the above description, the present teachings may also include a method of filling a sample substrate.  
      As mentioned above, the microcard may have other configurations including but not limited to the number of sample wells and reservoirs. A microcard  110  is depicted in  FIG. 4  in a closed position and is viewed facing an outer surface of plate  112 . Microcard  110  is similar in many respects to the microcard depicted in  FIG. 1 , but has  96  sample wells  120 . Sample wells  120  are each in fluid communication with a branch channel  126  to one of a plurality of main channels  122 . Channels  122  further communicate with reservoir  124 . Microcard  120  also comprises an area  170  where information about the card may be written or where a sticker containing information about the card or its contents may be affixed. Such information may be in the form of a bar code, written information, or any other form suitable for displaying desired characteristics of the card or the samples contained therein.  
      According to another embodiment, a microcard  210  is depicted in  FIG. 5 , which does not include a reservoir or feed channels, but is otherwise substantially similar to microcard  10 . Microcard  210  is depicted as having  96  sample wells  220 , but any number of sample wells may be provided. Microcard  210  also comprises a first plate  212  and a second plate  214  connected via a hinge  216 . Plate  214  includes closure elements  240  comprising a flexible annular rim  242  surrounding a cap  244 , which functions in a similar fashion to the closure element described above with reference to  FIGS. 1-3 . Microcard  210  may be used in a PCR environment whereby a user may desire to fill each sample well  220  separately with each of the reagent and the sample, or any other material desired to be tested. Microcard  210  may be suitable to have completely different reaction materials in one or more of sample wells  220 , as desired by a user, or it may be used in a situation where fill equipment such as a vacuum or centrifugal fill is not available. Test fluids may be introduced using a pipette, by hand or automatically, as well as by any other means suitable for filling a microcard sample well.  
      Once filled, microcard  210  may be closed in a similar fashion as described above and as depicted in  FIGS. 6A-6C , which show a partial section view of a sample well  220 . As seen in  FIG. 6A , sample well  220  has been filled or spotted with a desired sample  250  via, for example, pipetting. In this embodiment, sample  250  may comprise both the reagent and the sample. In addition, with this example, spotting may refer to the filling of either one or both of the reagent and the sample. Plate  214  is then positioned over plate  212  to a closed position as depicted in  FIG. 6B  and in a similar manner as described above in the embodiment of  FIGS. 1-3 . Because each well  220  is completely isolated within plate  212  a membrane may not be necessary to assist in isolating the various samples. Even though not required, it may be desirable, however, to include a membrane (not shown in  FIGS. 6A-6C ) similar to membrane  30  (see  FIGS. 3A-3D ) to assist in maintaining plates  212  and  214  in a closed relationship. Once closed, cap  244  may then be compressed to substantially seal sample well  220  in a similar fashion as described herein with flexible portions  220 - 1  and  246  bulging to compensate for displaced sample fluid as seen in  FIG. 6C .  
      According to another embodiment similar to microcard  210  depicted in  FIG. 5 , a closed microcard  310  is shown in  FIG. 7  having a first plate  312  and a second plate  314  and  96  sample wells  320 . Because the feed channels are not necessary in such a microcard, sample wells  320  may be offset and moved closer together to allow for a smaller overall microcard size and/or to allow for a higher sample well density within a microcard identical in size to microcard  210 . In other words, the sample wells in the  FIG. 7  embodiment are not positioned in a matrix, unlike the sample wells in the microcards shown in  FIGS. 1-6 .  
      In another exemplary embodiment,  FIG. 8  depicts a sample well  220  having an additional feature of a light pipe  280 . Although cap  244  is configured to be immersed within sample  250  to provide the benefit of minimizing the disadvantages of condensation within the well, light pipe  280  may be formed on or as part of flexible portion  246 . Light pipe  280  is designed to further extend within sample well  220  to further ensure that a portion of cap  244  is sufficiently immersed within the sample  250 . Light pipe  280  may be a cylindrical protrusion of polypropylene, or any other size or shape suitable for the desired radiation transmission characteristics desired with PCR testing. Light pipe  280  may also incorporate optics that may assist in focusing or directing radiation into and out of sample well  220 . Flexible annular rim  242  surrounds cap  244 , and functions in a manner similar to that described for  FIGS. 5-6 .  
      Although microcards  10 , 110 ,  210 , and  310  have been described above in relation to a card that has a first member and a second member movable with respect to one another, the present teachings could also apply to a card whereby the first and second members are fixed relative to one another. Such a card could be pre-spotted, as is done with conventional cards, but would contain a plurality of closure elements to substantially seal the sample wells. Essentially, a card of this configuration, instead of using a foil backing, could have a polypropylene member similar to the second member affixed to the first member and containing the closure elements. In this embodiment, for example, a pre-spotted card could incorporate closure elements, therefore allowing the staking to be replaced with moving closure elements in place to substantially seal the sample wells.  
      It will be apparent to those skilled in the art that various modifications and variations can be made to the structure and methods described above. Thus, it should be understood that the present teachings are not limited to the examples discussed in the specification. Rather, the present teachings are intended to cover modifications and variations.