Abstract:
Embodiments of the present invention, disclose a biological sample holder comprising a stake of biologically inert material, and a biological sample storage medium fixed to or retained on the stake by a retaining portion. This provides a means of holding a biological sample which is easy to handle and suitable for automation, for example in an array of such holders, allowing processing of multiple biological samples in parallel.

Description:
FIELD OF THE INVENTION 
       [0001]    The present invention relates to a biological sample holder and to a method of assembling a storage device. 
       BACKGROUND OF THE INVENTION 
       [0002]    Biological samples, such as blood samples taken for drug discovery and saliva taken for DNA profiling in criminal investigations, are typically held in an absorbent storage medium, which may comprise a membrane impregnated with chemicals for stabilising the sample. The samples are allowed to dry and, once dry, the biological storage medium can be transported to a testing facility for analysis. 
         [0003]    Typically, when the sample is to be tested, small pieces of the sample holding membrane are punched out. These pieces are small enough to minimise wasteful consumption of the sample but large enough to be handled and also to contain enough biological material for the test to be carried out successfully. 
         [0004]    Conventionally, this processing is done manually and samples are tested individually. However, greater demand for storage and extraction of genetic material has led to a requirement for greater throughput; the standard is now hundreds or thousands of extractions per day. Currently, matrix-based solutions for nucleic acid storage and extraction are limited in this respect because automated or multiple-sample processing of the samples is not compatible with the storage medium. 
         [0005]    It is an object of the present invention to mitigate the limitations associated with storing nucleic acid samples in matrices and provide a way to increase the speed and efficiency of sample processing. 
       SUMMARY OF THE INVENTION 
       [0006]    In accordance with a first aspect of the present invention, there is provided a biological sample holder comprising a stake of biologically inert material, and a biological sample storage medium fixed to or retained on the stake by a retaining portion of the stake, wherein, said medium includes a front surface and opposing rear surface, and wherein said retaining portion extends through the medium and beyond said front and rear surfaces. 
         [0007]    The retaining portion may comprise a recess in the stake, into which an edge of biological sample storage medium protrudes. This prevents excessive movement of the biological sample storage medium. 
         [0008]    The biological sample storage medium may comprise a membrane for absorbing liquid biological samples, which comprises a matrix disc. 
         [0009]    The biological sample storage medium may be made of a paper material. 
         [0010]    The stake may be tapered, optionally narrowing toward a distal end. 
         [0011]    The stake may be of plastics material. 
         [0012]    In accordance with a second aspect of the present invention, an array of biological sample holding devices supported on a base plate is provided. These stakes may each comprise biologically inert material, and may retain a biological sample storage medium thereon by means of a retaining portion of the stake, This provides the ability to process multiple biological samples in parallel. 
         [0013]    The base plate may comprise at least one asymmetric feature located on an outside edge. 
         [0014]    The base plate and the stake of the biological sample storage holder may be of a plastics material. 
         [0015]    The base plate may comprise a grid-coordinate system to identify each of the stakes. 
         [0016]    A gasket or seal may be provided on the base plate around each of the stakes that reduces or prevents evaporation and/or spillage losses during processing of the biological sample. 
         [0017]    The base plate may comprise a computer readable tag to provide reliable identification of the samples. 
         [0018]    In accordance with a third aspect of the present invention, there is provided an apparatus for storing and processing in parallel plural biological samples, the apparatus comprising: 
         [0019]    an array of biological sample holders; and 
         [0020]    a tray comprising an array of wells; 
         [0021]    wherein the position of the wells corresponds to the position of the devices in said array, and the depth and diameter of the wells exceeds the length and diameter of the stakes in said array. 
         [0022]    The tray may be a polymerase chain reaction (PCR) tray. 
         [0023]    In accordance with a fourth aspect of the present invention, there is provided a method of assembling a device for storing and processing at least one biological sample, the method comprising: 
         [0024]    providing one or more stakes; and 
         [0025]    fixing or retaining one or more biological sample storage medium/media onto the stake(s) such that the or each stake at least partly penetrates a first surface of its associated biological sample storage medium and emerges from an opposing side of said associated medium. 
         [0026]    The method may comprise providing a plurality of stakes arranged in a two dimensional array on a base plate. 
         [0027]    The base plate and stakes may be made of a plastics material and may be injection moulded as a single component. This has the advantage that the component may be fabricated in a simple single step. Alternatively, the base plate and the or each stake may be manufactured as discrete parts that are fixed together. This has the advantage that the design of the array is flexible. A gasket or seal may be provided around the or each stake. 
         [0028]    The biological sample storage medium may be fixed to or retained on said stake by a heat staking process if the stake is of plastics, or retained on the stake by an interference fit. 
         [0029]    The method may also comprise, in any suitable order, forming a biological sample storage medium from a matrix; fixing it to, or retaining it on a stake; and applying a biological sample to the biological sample storage medium. 
         [0030]    A further aspect of the invention provides the use of the holder of the first aspect, the array of the second aspect or the apparatus of the third aspect for storing and/or processing a biological sample. 
         [0031]    Further features and advantages of the invention will become apparent from the following description of illustrative embodiments of the invention, given by way of example only, which is made with reference to the accompanying drawings. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0032]      FIG. 1   a  shows a perspective exploded view of a device for holding a disc of biological sample holding membrane according to a first embodiment of the present invention; 
           [0033]      FIG. 1   b  shows a perspective view of a device for holding a disc of biological sample holding membrane according to the first embodiment of the present invention; 
           [0034]      FIG. 2   a  shows a longitudinal cross-section view of a biological sample holder according to a second embodiment of the present invention; 
           [0035]      FIG. 2   b  shows longitudinal cross-section views of a biological sample holder with a retainer formed by a heat-staking process according to a third embodiment of the present invention; 
           [0036]      FIG. 3   a  shows a perspective view of an array of biological sample holders according to a fourth embodiment of the present invention; 
           [0037]      FIG. 3   b  shows an exploded perspective view of an array of biological sample holders according to a fifth embodiment of the present invention; 
           [0038]      FIG. 3   c  shows an exploded perspective view of a PCR tray corresponding with an array of biological sample holders according to a sixth embodiment of the present invention; 
           [0039]      FIG. 3   d  shows a cross-section view of a PCR well according to the sixth embodiment of the present invention; 
           [0040]      FIG. 4  shows a cross-section view of the edge of the base plate of an array of devices for holding discs of biological sample holding membrane according to the seventh embodiment of the present invention; 
           [0041]      FIG. 5  shows a perspective view of an array of devices for holding discs of biological sample holding membrane comprising a gasket according to an eighth embodiment of the present invention; 
           [0042]      FIG. 6  shows a plan view of the base plate of an array of devices for holding discs of biological sample holding membrane comprising a grid-coordinate reference system according to a ninth embodiment of the present invention; 
           [0043]      FIG. 7  shows a side aspect of an array of devices for holding discs of biological sample holding membrane comprising a computer readable tag coded with identification data according to a tenth embodiment of the present invention; 
           [0044]      FIG. 8  shows a plan view of an array of devices for holding discs of biological sample holding membrane, wherein the base plate of the array comprises an asymmetric external geometry according to an eleventh embodiment of the present invention. 
       
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
       [0045]      FIG. 1   a  shows an exploded perspective view of a biological sample holder  10  according to an embodiment of the present invention.  FIG. 1   b  shows a perspective view of a biological sample holder  10  in assembled form according to the embodiment. Longitudinal and transverse axes are defined by the arrows in  FIGS. 1   a  and  1   b  and labelled L and T respectively. The biological sample holder  10  comprises a stake  11  having a base portion  12 , a shaft portion  13 , a tip portion  14  and a retainer or retaining portion  15  which fixes or retains, on the stake  11 , a piece of a biological sample holding medium. Typically, the stake  11  is 20 mm long and 2 mm in diameter at its base portion  12 . The stake  11  may be solid or hollow. In this example, the medium is a membrane (referred to hereinafter as a matrix disc  16 ). The matrix disc  16  comprises a substantially flat, circular disc of matrix material having a opposing front  16   a  and rear  16   b  surfaces, suitable for holding wet or dried biological material, with a hole  17  substantially in the centre of the matrix disc  16 . The stake  11  extends through the matrix disc  16  beyond the front and rear surfaces  16   a  and  16   b . The matrix disc  16  may be made of a paper material, such as FTA® elute paper. Typically, the matrix disc  16  and hole  17  are formed by punching the matrix disc  16  from a larger sheet of membrane. The matrix disc  16  is typically 3 mm in diameter and 1 mm thick. The hole  17  is typically 1 mm in diameter and may be designed such that it provides an interference fit with the tip portion  14 . The hole  17  may be cut by the tip portion  14  of the stake  11 . 
         [0046]    In an embodiment, the biological sample holder  10  is substantially circular in transverse cross-section, though it will be apparent to one skilled in the art that embodiments with other cross-sectional profiles are also possible. 
         [0047]    The tip portion  14  of the biological sample holder  10  is inserted into the hole  17  in the centre of the matrix disc  16 . In an embodiment, the stake is tapered such that the base portion  12  has a larger diameter than the tip portion  14  in order to prevent excessive movement of the matrix disc  16  along the shaft portion  13  toward the base portion  12  during assembly. The additional diameter of the base portion  12  also provides an increase in overall structural strength, to minimise the chance of damage to the shaft, while the diameter of the tip portion  14  is still small enough to be accommodated by the hole  17  in the matrix disc  16 . The biological sample holder  10  may have any suitable longitudinal cross sectional shape. It may be cylindrical, tapered, or stepped from the base portion  12  to the tip portion  14 . 
         [0048]      FIG. 2   a  shows an embodiment of the present invention wherein the retainer  15  of the biological sample holder  10  comprises features to prevent excessive movement of the matrix disc  16  along the longitudinal axis of the shaft portion  12  such as to prevent the matrix disc  16  from moving toward the base portion  12  or from being completely removed from the biological sample holder  10 . The longitudinal cross-sectional profile of the shaft may comprise one or more protrusions  21  designed to prevent movement of the matrix disc  16 . 
         [0049]    The protrusions  21  may be formed around part of, or the entire circumference of the stake  11 . In order for the matrix disc  16  to be fitted to the stake  11 , the matrix material from which the matrix disc  16  is made may be flexible such that the hole  17  can expand to pass the outermost protrusion  21  and, once past the outermost protrusion  21 , contract to lie within the recess formed between the outermost and innermost protrusions  21 . In another embodiment according to the present invention, as shown in  FIG. 2   b , the retainer  15  comprises a tip portion  14  which has a smaller diameter than the shaft portion  12  such that a shoulder  22  is formed at the interface between the shaft portion  12  and the tip portion  14 . The tip portion  14  is deformed, e.g. by the application of heat and pressure to a stake  11  of plastics material, in a heat staking process to trap the matrix disc  16  between the shoulder  22  and the deformed tip portion  23  and thus prevent movement along the longitudinal axis of the biological sample holder  10 . 
         [0050]    The biological sample holder  10  may be made from any chemically and biologically inert material that can withstand a temperature of a PCR reaction-currently around 95° C., although higher or lower temperatures could be employed. Suitable materials include polycarbonate, polystyrene, or polypropylene. The biological sample holder  10  must be rigid and strong enough to withstand the forces applied to it when mounting the matrix disc  16  on to the tip portion  14 . 
         [0051]    In a further embodiment according to the present invention, depicted in  FIG. 3   a , a plurality of biological sample holders  10  are arranged to form an array  30  such that multiple samples may be processed in a single step. The holders may be as shown and described with reference to  FIG. 1 ,  2   a  or  2   b.    
         [0052]    Each of the biological sample holders  10  are physically connected at the base portion  12  to a base plate  31 . 
         [0053]    In a preferred embodiment the array  30  of biological sample holders  10  and the base plate  31  are manufactured by injection moulding as a single component. In this embodiment, it may be particularly advantageous to form the stakes  11  without the protrusions  21 , since it is difficult to form a two-dimensional array of stakes  11  having these protrusions as part of a single injection mould. Accordingly, in this embodiment, it may be advantageous to form stakes  11  having a reduced tip diameter, and use the heat-staking method to form the retainer  15 , as described above in relation to  FIG. 2   b.    
         [0054]    Other methods of manufacture are envisaged; for example, the biological sample holders  10  and the base plate  31  may be manufactured as separate parts as shown in  FIG. 3   b . This allows greater flexibility in the design of the array  30  and, in particular, in the number of biological samples holders  10  that form the array  30 . The biological sample holders  10  may be moulded individually or in rows  32 , for example, and the base plate may simply be formed with an array of holes  33  into which the base portions  11  of the biological sample holders  10  sit. The biological sample holders  10  may be fixed to the base plate  31  by an ultrasonic weld or any other suitable method. 
         [0055]    The dimensions of the base plate  31  and the positions of the biological sample holders  10  on the base plate  31  are chosen to correspond with the dimensions and positions of wells in a tray of wells. In this example the tray is a polymerase chain reaction (PCR) type tray  34  and the positions of wells  35  within the PCR tray  34  may be as shown in  FIG. 3   c . Typically, the wells  35  in the PCR tray  34  will be at least partially filled with an elution liquid  36  designed to elute the nucleic acid. In the arrangement shown in  FIGS. 3   a ,  3   b  and  3   c , the biological sample holders  10  form an 8×12 rectangular array, however, it will be appreciated that any other one- or two-dimensional arrangement corresponding to an equivalent arrangement of wells  35  in a PCR tray  34  is possible. In a preferred embodiment according to the present invention, the external dimensions of the base plate  31  correspond to the SBS standard laboratory footprint such that the array  30  may be handled by standard laboratory material handling equipment. Typically, the base plate is 127.76 mm long and 85.48 mm wide. The height of the biological sample holder  10  is designed such that when the base plate  31  of the array  30  is attached to the PCR tray  34 , the tip portion  14  of the biological sample holders  10  reaches far enough into to the well  35  that the matrix disc  16  may be immersed in elution liquid  36  without the tip portion  14  or the matrix disc  16  coming into physical contact with the internal walls of the PCR well  35 . This is illustrated in  FIG. 3   d . There is also a design restriction on the diameter of the matrix disc  16  to be smaller than the diameter of the PCR well  35 , and for the central longitudinal axis of the biological sample holder  10  to be substantially parallel to the central longitudinal axis of the PCR well  35 . 
         [0056]      FIG. 4  shows, in cross-section, a further embodiment of the present invention in which the external edges of the base plate  31  of the array  30  form a mechanical clip  41  to hold the array  30  in place on the PCR tray  34 . The edge of the PCR tray  34  is shown in contact with the base plate  31  of the array  30 . 
         [0057]    The PCR tray  34  has a corresponding protrusion  42  at its edge, which binds with the clip  41 . The dimensions of the clip  41  are such that it is flexible enough that the base plate  31  may be attached to and removed from the PCR tray  34  by application of appropriately directed forces, but stiff enough that there is negligible movement of the base plate  31  relative to the PCR tray  34  when the two components are connected and such that the base plate  31  cannot detach from the PCR tray  34  inadvertently when the combined components are gripped solely by the edges of the base plate  31 , either by a human operator or a mechanical handling system. 
         [0058]    In a further embodiment according to the present invention, as shown in  FIG. 5 , the array  30  comprises a gasket  51  fixed to the base plate  31  and surrounding each of the biological sample holders  10 . The gasket  51  is manufactured from an impermeable material and is of sufficient thickness and flexibility to form a suitable seal between the base plate  31  of the array  30  and the PCR tray  34  to minimise loss of the elution liquid  36  by evaporation or spillage. Rather than the continuous gasket  51 , individual seals may be employed for each stake, for example ‘O’ ring seals (not shown). 
         [0059]      FIG. 6  shows an embodiment according to the present invention wherein the base plate  31  comprises identifying grid-coordinates  61  correlating with the positions of each of the biological sample holders  10 , to enable identification and addressing of individual samples. 
         [0060]      FIG. 7  shows a further embodiment according to the present invention wherein the base plate  31  comprises a identification tag  71  comprising coded computer readable identification information. In an embodiment, the tag  71  comprises a barcode  72  that can be scanned and compared with a database of sample identification codes; other types of tag may be used, for example an RFID tag. 
         [0061]    In a further embodiment according to the present invention, as shown in  FIG. 8 , the base plate  31  comprises an asymmetric geometry such that the array  30  can only couple with the PCR tray  34  in a single orientation. In the embodiment shown, this is achieved by incorporating a bevel  81  in one of the four corners of the base plate  31  and corresponding PCR tray  34 . However, it will be apparent that other geometries will also achieve the same result. 
         [0062]    The biological sample holders  10  and the array  30  described by the above embodiments may be used in any process whereby a liquid sample is stored in dried form within a matrix material and then subsequently removed from the matrix material by elution. A typical process compatible with DNA amplification techniques may involve the following steps: inserting the one or more biological sample holders  10  in the wells of a tray with each well having a volume of at least 800 μL so that each matrix is immersed in 500 μL of water. Transferring the one or more biological sample holders  10  to a PCR tray  34  that contains 30 μL of water in each well  35 ; transferring the array  30  and PCR tray  34 , together, to a thermal cycler and heating to 95° C. for thirty minutes; pulse vortexing the array  30  and PCR tray  34 , together, sixty times; spinning the array  30  and PCR tray  34 , together, in a centrifuge for thirty seconds at 1000×g; removing the array  30  from the PCR tray  34 ; and passing the PCR tray  34  on to be analysed. 
         [0063]    The above embodiments are to be understood as illustrative examples of the invention. Further embodiments of the invention are envisaged. For example, in some embodiments the shoulder portion  22  of  FIG. 2   b  is omitted and the shaft portion  13  tapered, in which case the retainer  15  holding the matrix disc  16  comprises the taper of the stake  11  and the deformed tip portion  23 . It is to be understood that any feature described in relation to any one embodiment may be used alone, or in combination with other features described, and may also be used in combination with one or more features of any other of the embodiments, or any combination of any other of the embodiments. Furthermore, equivalents and modifications not described above may also be employed without departing from the scope of the invention, which is defined in the accompanying claims.