Filter plate

Abstract

A filter plate (10) is used by being attached to a sample container having a sample-accommodating part of which an upper part is opened, the filter plate (10) comprising: a base material (11, 31) provided with first through-holes (112, 312) for circulating a sample liquid at positions corresponding to the sample-accommodating part; first filters (12, 32) provided in the first through-holes; and support bodies (13, 33) which are disposed adjacent to the side of the first filter from which the sample liquid flows out, and in which second through-holes larger than the pore size of the first filters are formed. One aspect of the filter plate (10) is used for a microplate having a plurality of wells. The first through-holes are provided in the base material at positions corresponding to the wells of the microplate. The first filters are sheet-form filters disposed on one surface of the base body so as to cover all of the first through-holes.

Description

filter plate

[0001] The present invention relates to a filter plate that is attached to a sample container such as a microplate to filter a sample liquid.

[0002] Conventionally, when a sample liquid, such as a biological sample liquid, is poured into a well of a microplate, a filter is used to remove impurities from the sample liquid and extract a target substance from the sample liquid. Because the pore size of such a filter is small, the sample liquid often does not pass through the filter simply by dropping the sample liquid onto the filter. Therefore, a procedure known as centrifugal filtration is performed in which the sample liquid is dropped onto a filter provided corresponding to each well, and then centrifugal force is applied using a centrifuge to draw the sample liquid into the well (e.g., Patent Documents 1 and 2).

[0003] Special table 2020-519440 Publication Special table 2007-509633

[0004] When centrifugal filtering a sample liquid derived from a living organism, a thin filter with a small pore size is often used, which has the problem that the filter is easily broken when subjected to operations that apply force to the filter, such as centrifugal filtration.

[0005] The problem that the present invention aims to solve is to provide a filter plate that can prevent the filter from breaking when performing an operation in which force is applied to the filter, such as centrifugal filtration of a sample liquid.

[0006] The present invention, which has been made to solve the above-mentioned problems, is a filter plate that is attached to a sample container having a sample holding section that is open at the top, and is characterized by comprising: a) a base material having a first through-hole formed at a position corresponding to the sample holding section, through which sample liquid flows; b) a first filter provided in the first through-hole; and c) a support body that is positioned adjacent to the side of the first filter from which the sample liquid flows out, and that has a second through-hole formed therein that is larger in pore size than the first filter.

[0007] The filter plate according to the present invention is fixed to a sample container so that the position of the opening of the sample container's sample storage portion coincides with the position of the first through-hole in the substrate, and the support is located closer to the sample container than the filter. In the filter plate according to the present invention, a first filter used for centrifugal filtration or the like is provided in the first through-hole in the substrate fixed to the sample container. The filter plate according to the present invention further includes a support disposed adjacent to the side of the first filter from which the sample liquid flows out. Therefore, when performing operations that apply force to the filter, such as centrifugal filtration, the filter is supported from the back side (the side facing the sample container), thereby preventing damage to the first filter. Furthermore, the support is formed with second through-holes larger than the pore size of the first filter used in centrifugal filtration, so that the sample liquid that has passed through the filter is not prevented from being introduced into the sample container.

[0008] In the filter plate of the present invention, it is preferable that the filter plate is used in a microplate having a plurality of wells, the first through-holes are provided in the base material at positions corresponding to each well of the microplate, and the first filter is a sheet-like filter arranged on one side of the base material so as to cover all of the first through-holes.

[0009] The filter plate according to the present invention can be used in a microplate having multiple wells, as in the above-described embodiment. In this case, the first filter can be easily attached by using a sheet-like filter disposed on one surface of the base so as to cover all of the first through-holes, as in the above-described embodiment.

[0010] In the filter plate according to the present invention, the support is preferably a second filter formed by laminating an adhesive sheet to a sheet-like material, with the second through-holes formed therein.

[0011] In the filter plate of the above embodiment, the first filter and the second filter are adhered to each other by the adhesive sheet of the support, thereby preventing the sample liquid from flowing between them.

[0012] It is preferable that the filter plate according to the present invention further comprises a flexible plate-shaped packing material that is arranged on the side of the support from which the sample liquid flows out and has third through holes formed at positions corresponding to each of the plurality of wells.

[0013] In the filter plate of the above aspect, the flowing sample liquid can be introduced from the first through-hole of the substrate into the well corresponding to the through-hole without leakage.

[0014] By using the filter plate according to the present invention, damage to the filter can be prevented when a force is applied to the filter, such as centrifugal filtration of a sample liquid.

[0015] 1A is a top view (a), a vertical cross-sectional view (aa), and a bottom view (c) showing a first embodiment of a filter plate according to the present invention; FIG. 1B is a vertical cross-sectional view showing a filter and packing seal including a filter and packing, which is attached to a substrate during the manufacture of the filter plate of the first embodiment; FIG. 2 is an exploded perspective view of a filter and packing seal according to the first embodiment; FIG. 3 is a vertical cross-sectional view showing the filter plate of the first embodiment attached to a microplate; FIG. 4 is a schematic view showing the microplate and the filter plate of the first embodiment attached to a centrifuge; FIG. 1A is a top view (a), a vertical cross-sectional view (b) and a bottom view (c) showing a second embodiment of a filter plate according to the present invention; FIG. 1A is a partial enlarged vertical cross-sectional view (a) and a partial enlarged bottom view (b) of the filter plate of the second embodiment; FIG. 1B is a partial enlarged vertical cross-sectional view showing the filter plate of the second embodiment attached to a microplate; FIG. 1C is a partial vertical cross-sectional view showing a filter plate of a modified example of the first embodiment; FIG. 2A is a vertical cross-sectional view of a filter plate according to an embodiment related to the present invention. A longitudinal cross-sectional view showing a filter and packing seal including a filter and packing that is attached to a base material during manufacturing of a filter plate of a related embodiment. An exploded perspective view of a filter and packing seal of a related embodiment. A partial enlarged view of a second filter sheet of a related embodiment. A partial enlarged view showing a state in which a pipette tip is inserted into one of the first through-holes and pressed against the second filter sheet to extract sample liquid in a related embodiment.

[0016] An embodiment of a filter plate according to the present invention will be described with reference to FIGS.

[0017] (1) First Embodiment Fig. 1 shows a filter plate 10 according to a first embodiment. Fig. 1(a) is a top view of the filter plate 10, Fig. 1(b) is a longitudinal cross-sectional view of the filter plate 10 taken along line aa, and Fig. 1(c) is a bottom view of the filter plate 10. The filter plate 10 includes a substrate 11, a first filter sheet 12, a second filter sheet 13, and a packing material 14.

[0018] The substrate 11 includes a plastic plate-like member 111 having a rectangular planar shape and a plurality of first through holes 112 having a circular planar shape provided in the plate-like member 111. The first through holes 112 are arranged in a total of 96, with 12 arranged parallel to the long sides of the rectangle and 8 arranged parallel to the short sides. The arrangement of the first through holes 112 corresponds to the arrangement of the wells of a 96-well microplate to which the filter plate 10 is attached. On the top surface of the substrate 11, the ends of the long sides are marked with numbers 1 to 12 corresponding to the columns in which the first through holes 112 are arranged, and the ends of the short sides are marked with eight letters A to H corresponding to the rows in which the first through holes 112 are arranged. These numbers and letters are symbols for identifying each first through hole 112.

[0019] Of the four corners of the rectangular base material 11 (plate-shaped member 111), two adjacent corners across one short side are formed with C-faces 113 cut off at an angle of 45° with respect to the short side. By providing the C-faces 113, the orientation of the filter plate 10 can be easily identified.

[0020] A rectangular frame-shaped surrounding wall 114 made of the same material as the substrate 11 extends downward from the outer periphery of the lower surface of the substrate 11. In this embodiment, the surrounding wall 114 is provided perpendicular (vertical) to the surfaces of the first filter sheet 12 and the second filter sheet 13, but the surrounding wall 114 is not limited to being provided vertically as long as it is provided to surround the microplate insertion space 115.

[0021] The area below the bottom surface of the substrate 11 and surrounded by the surrounding wall 114 forms a microplate insertion space 115 into which a 96-well microplate is inserted. By inserting a microplate into the microplate insertion space 115, the microplate and the substrate 11 are positioned and fixed relative to each other. The microplate insertion space 115 is basically a rectangular parallelepiped, but a corner portion 116 (see FIG. 1(c)) made of the same material as the surrounding wall 114 is provided directly below the portion of the rectangular parallelepiped where the substrate 11 is cut off at the C-face 113. In other words, the planar shape of the microplate insertion space 115 and the planar shape of the top surface of the substrate 11 are substantially the same.

[0022] The first filter sheet 12 is made of a filter material made of nylon 66 and having a mesh size (filtration particle size) of 30 μm. The first filter sheet 12 has approximately the same shape as the area of ​​the underside of the substrate 11 that is surrounded by the surrounding wall 114, and is attached to the underside of the substrate 11 so as to cover that area. The portion of the first filter sheet 12 that faces the first through-holes 112 functions as the filter in the present invention.

[0023] The second filter sheet 13 is a sheet material in which an adhesive sheet 132 is bonded to one surface of a polyethylene terephthalate (PET) sheet 131 (one adhesive surface of a double-sided adhesive sheet is bonded to the PET sheet 131) and multiple through-holes are formed by irradiating the sheet with laser light (the through-holes are schematically illustrated only in FIG. 3 , and are not illustrated in FIGS. 1 and 2 ). The second filter sheet 13 also has approximately the same shape as the area surrounded by the surrounding wall 114 on the underside of the substrate 11 and is attached to cover the underside of the first filter sheet 12. The second filter sheet 13 functions as a support in the present invention. The through-holes provided in the second filter sheet 13 correspond to the second through-holes in the present invention. The second filter sheet 13 has through-holes formed therein that are larger than the pore size of the first filter sheet 12 but smaller than the diameter of the first through-holes 112.

[0024] If the through-holes of the second filter sheet 13 are too small, the sample liquid that has passed through the first filter sheet 12 may not pass through the second filter sheet 13 easily. Furthermore, if the through-holes of the second filter sheet 13 are too large, the area of ​​contact with the first filter sheet 12 will be small, resulting in a narrower region for supporting the first filter sheet 12. Taking these factors into consideration, the diameter of the through-holes formed in the second filter sheet 13 is preferably 0.1 μm or more and 1 mm or less. Furthermore, considering that the second filter sheet 13 can be used regardless of the pore size of the first filter sheet 12, a diameter of 1 μm or more and 500 μm or less is more preferable. This allows the sample liquid that has passed through the first filter sheet 12 to pass through smoothly and more reliably supports the first filter sheet 12, regardless of the pore size of the first filter sheet 12.

[0025] The packing material 14 is made of a plate-like member 141 that is thicker than the first filter sheet 12 and the second filter sheet 13, and that has third through holes 142 formed in positions corresponding to the first through holes 112 of the base material 11. The packing material 14 has substantially the same planar shape as the first filter sheet 12 and the second filter sheet 13, and is provided on the underside of the base material 11 so as to sandwich the first filter sheet 12 and the second filter sheet 13 therebetween. The plate-like member 141 of the packing material 14 is made of silicone rubber and is flexible.

[0026] As shown in Figures 2 and 3, the second filter sheet 13 and the packing material 14 are bonded to each other by a first double-sided adhesive film 15 having a hole formed at a position corresponding to the third through-hole 142 of the packing material 14 (the first double-sided adhesive film 15 is not shown in Figure 1). Note that Figure 2 uses different horizontal and vertical scales (i.e., the vertical direction is longer) to clearly show each component. The first filter sheet 12 and the second filter sheet 13 are bonded to each other by an adhesive sheet 132 located on the upper surface of the second filter sheet 13. The first filter sheet 12 and the base material 11 are bonded to each other by a second double-sided adhesive film 16 having a hole formed at a position corresponding to the first through-hole 112 of the base material 11 (the second double-sided adhesive film 16 is not shown in Figure 1). The first double-sided adhesive film 15 and the second double-sided adhesive film 16 each have the same planar shape as the first filter sheet 12, the second filter sheet 13, and the packing material 14. That is, two notches 123, 133, 143, 153, 163 are provided for each of the first filter sheet 12, the second filter sheet 13, the packing material 14, the first double-sided adhesive film 15, and the second double-sided adhesive film 16 at locations corresponding to the two C-faces of the base material 11 (see Figure 3).

[0027] When manufacturing the filter plate 10, first, the packing material 14, the first double-sided adhesive film 15, the second filter sheet 13, the first filter sheet 12, and the second double-sided adhesive film 16 are stacked and bonded together, starting from the bottom, by aligning the respective cutouts 143, 153, 133, 123, and 163. The second filter sheet 13 is positioned with the adhesive sheet 132 facing upward. Next, this integrated assembly (referred to as the "filter / packing seal 20") is fitted into the space (microplate insertion space 115) surrounded by the surrounding wall 114 on the underside of the substrate 11, aligning the cutout with the corner 116 of the substrate 11, and then attached to the underside of the substrate 11 with the second double-sided adhesive film 16. This completes the filter plate 10.

[0028] A release paper may be attached to the surface of the adhesive sheet 132 (the surface on which the first filter sheet 12 is attached), and the release paper may remain attached to the adhesive sheet 132 during the manufacture of the filter plate 10, and then peeled off from the adhesive sheet 132 during the manufacture of the filter / packing seal 20. Alternatively, a release paper may be attached to the back surface of the packing material 14 (the surface on which the microplate is attached), and the release paper may remain attached to the packing material 14 during the manufacture of the filter / packing seal 20 and the filter plate 10, and then peeled off from the packing material 14 immediately before the filter plate 10 is attached to the microplate. Covering the surface of the adhesive sheet 132 and the back surface of the packing material 14 with the release paper can prevent dust from adhering to the surface of the adhesive sheet 132 and the back surface of the packing material 14 during the manufacture of the filter plate 10, etc. Furthermore, it can prevent dust from getting into the wells of the microplate.

[0029] Hereinafter, a method of using the filter plate 10 of the first embodiment will be described with reference to FIGS.

[0030] First, a 96-well microplate 90 is prepared, the shape of which corresponds to the microplate insertion space 115 of the filter plate 10. Then, the C-face of the microplate 90 is aligned with the corners 116 of the filter plate 10, and the microplate 90 is inserted from below into the microplate insertion space 115 of the filter plate 10 ( FIG. 4 ). Next, the microplate 90 and the filter plate 10 are pressed against each other. At this time, because the packing material 14 located between the microplate 90 and the substrate 11 is flexible, the microplate 90 and the filter plate 10 are pressed against each other, and are thus connected liquid-tightly without any gaps.

[0031] Next, with the filter plate 10 fixed to the microplate 90, a predetermined amount of sample liquid 80 is dropped into each of the first through-holes 112 of the substrate 11 using a micropipette (not shown). At this time, it is sufficient that the dropped sample liquid 80 rests on the filter; there is no need to press the tip of the tip attached to the nozzle of the micropipette firmly against the surface of the first filter sheet 12. Therefore, it is not necessary to insert the tip deeply into the first through-hole 112. For example, it is possible to easily drop sample liquid 80 into multiple first through-holes 112 simultaneously using a multi-channel micropipette with multiple nozzles.

[0032] After dropping the sample liquid 80 into the first through-holes 112 of the filter plate 10, the microplate 90 and the filter plate 10 are tilted nearly vertically and set in the holder 701 of the centrifuge 70 (see FIG. 5 ). To maintain close contact between the microplate 90 and the filter plate 10 set in the holder 701 of the centrifuge 70, they may be fastened together with a rubber band, clips, or the like. The centrifuge 70 is then driven to rotate the holder 701 at high speed around the vertically extending rotation axis 71. This presses the sample liquid 80 in the first through-holes 112 of the filter plate 10 against the first filter sheet 12, removing any particles (such as debris) larger than the pore size of the first filter sheet 12 and filtering the sample liquid into the wells 91 of the microplate 90.

[0033] Filter plates come in a variety of pore sizes, and different filter plates are used depending on the intended use. For example, when centrifugal filtering of biological sample liquids or capturing intracellular substances (such as nucleic acids), thin filters with small pore sizes are often used. Because filter plates with small pore sizes are thin and weak, conventional filter plates can break during centrifugal filtering.

[0034] In contrast, in the filter plate 10 of this embodiment, a second filter sheet 13 is disposed adjacent to the underside (the side from which the sample liquid flows out) of a first filter sheet 12 used for the purpose of filtering the sample. Furthermore, the second filter sheet 13 is not mesh-like, but is a sheet material with multiple through-holes formed therein, and contacts the first filter sheet 12 at a surface rather than at points or lines. Therefore, during centrifugal filtration, the first filter sheet 12 is supported by the second filter sheet 13 from its back side (the side facing the microplate 90), thereby preventing damage to the first filter sheet 12. Furthermore, because the second filter sheet 13 has through-holes larger than the pore size of the first filter sheet, the sample liquid that has passed through the first filter sheet 12 is not prevented from being drawn into the wells 91 of the microplate 90.

[0035] Furthermore, because the filter plate 10 and the microplate 90 are connected in a liquid-tight state by the flexible packing material 14, when the sample liquid 80 in the first through-holes 112 is drawn into the wells 91 by centrifugal force, the sample liquid is prevented from flowing into other wells 91. This makes it possible to prevent different sample liquids from being supplied to the wells 91 in a mixed state.

[0036] (2) Second Embodiment Figures 6 and 7 show a filter plate 30 according to a second embodiment. Figure 6(a) is a top view of the filter plate 30, Figure 6(b) is a vertical cross-sectional view of the filter plate 30 taken along line bb, and Figure 6(c) is a bottom view of the filter plate 30. Figure 7(a) is a partially enlarged view of Figure 6(b), and Figure 7(b) is a partially enlarged view of Figure 6(c).

[0037] The filter plate 30 includes a base material 31 , a filter sheet 32 ​​, a reinforcing sheet material 33 , a packing material 34 , and a cylindrical body 37 .

[0038] The substrate 31 is made of a plastic plate-like member having a rectangular planar shape, and has 24 first through-holes 312 arranged parallel to the long sides and 16 first through-holes 312 arranged parallel to the short sides, for a total of 384 first through-holes 312 having a circular planar shape. The arrangement of the first through-holes 312 corresponds to the arrangement of the wells of the 384-well microplate to which the filter plate 30 is attached.

[0039] The substrate 31 is made up of a first substrate 3111 and a second substrate 3112 attached to the lower part of the first substrate 3111. A rectangular frame-shaped surrounding wall 3114 extends downward from the outer periphery of the lower surface of the first substrate 3111. In this embodiment, the surrounding wall 3114 is also provided perpendicular (vertical) to the surfaces of the filter sheet 32 ​​and the reinforcing sheet material 33, but the surrounding wall 3114 is not limited to being provided in the vertical direction as long as it is provided to surround the microplate insertion space 115.

[0040] The filter sheet 32, the reinforcing sheet material 33, and the second base material 3112 are attached in this order from the first base material 3111 side to the portion surrounded by the surrounding wall 3114 on the underside of the first base material 3111. The length of the surrounding wall 3114 is greater than the combined thickness of the filter sheet 32, the reinforcing sheet material 33, and the second base material 3112, and the space below the second base material 3112 and surrounded by the surrounding wall 3114 forms the microplate insertion space 315. The first base material 3111 and the second base material 3112 each have 384 through holes 3121, 3122, and the first through holes 312 are composed of the through holes 3121, 3122 that are positioned correspondingly when the first base material 3111 and the second base material 3112 are stacked one on top of the other. The through-hole 3121 is cylindrical and has a uniform inner diameter in the depth direction (the vertical direction in FIG. 7A), whereas the through-hole 3122 has a tapered shape with an inner diameter that is smaller at the bottom than at the top.

[0041] A cylinder 37 is provided on the underside of the second base material 3112 so as to extend downward from the periphery of the first through-hole 312. The cylinder 37 is made of the same plastic as the second base material 3112 and is molded integrally with the second base material 3112. The outer diameter of the cylinder 37 is slightly smaller than the inner diameter of the wells of a 384-well microplate, and the inner diameter of the cylinder 37 is equal to the inner diameter of the through-hole 3122 on the underside of the second base material 3112. The length of the cylinder 37 is shorter than the depth of the wells of a 384-well microplate.

[0042] The filter sheet 32 ​​and the reinforcing sheet material 33 are interposed between the first base material 3111 and the second base material 3112. The material and mesh size of the filter sheet 32 ​​are the same as those of the first filter sheet 12 of the first embodiment. The reinforcing sheet material 33 (corresponding to the support in the present invention) is, for example, a sheet material in which an adhesive sheet is bonded to the surface of a sheet made of a resin that is harder than the silicone rubber that constitutes the packing material 34 described below, and similar to the second filter sheet 13 in the first embodiment, a large number of through holes (corresponding to the second through holes in the present invention) are formed therein, each having a diameter larger than the pore size of the filter sheet 32 ​​and smaller than the diameter of the first through holes 312.

[0043] Because the reinforcing sheet material 33 has through holes with a diameter larger than the pore size of the filter sheet 32, the reinforcing sheet material 33 does not impede the flow of sample liquid that has passed through the filter sheet 32. Furthermore, because the reinforcing sheet material 33 has through holes with a diameter smaller than the diameter of the first through holes 312, the filter sheet 32 ​​can be supported inside the first through holes 312 regardless of the positions at which the through holes are formed. In the second embodiment, for the reasons described in the first embodiment, the diameter of the through holes is preferably 0.1 μm or more and 1 mm or less, and more preferably 1 μm or more and 500 μm or less. As in the first embodiment, the filter sheet 32 ​​and the reinforcing sheet material 33 cover the entire upper surface of the second base material 3112, and the portion of the filter sheet 32 ​​located within the first through holes 312 functions as a filter in the present invention.

[0044] The packing material 34 is made of a plate-like member made of silicone rubber, and is attached to the second base material 3112 so as to cover the lower surface of the second base material 3112 except for the cylindrical body 37. Therefore, the packing material 34 has a third through-hole 342 corresponding to the cylindrical body 37.

[0045] Although detailed explanation and illustration are omitted, the first substrate 3111 and the filter sheet 32, the reinforcing sheet material 33 and the second substrate 3112, and the second substrate 3112 and the packing material 34 are bonded together with an adhesive or an adhesive sheet with adhesive applied to both sides. The filter sheet 32 ​​and the reinforcing sheet material 33 are bonded together with an adhesive sheet provided on the reinforcing sheet material 33.

[0046] The filter plate 30 is used as follows. First, a microplate 90A (384 holes) having 384 wells 91A is prepared. Then, as shown in FIG. 8 , the microplate 90A is inserted from below into the microplate insertion space 315 of the filter plate 30, ensuring that the corresponding cylinder 37 is inserted into each well 91A of the microplate 90A. Next, the microplate 90A and the filter plate 30 are pressed against each other. By pressing the microplate 90A and the filter plate 30 together in this manner, the flexible packing material 34 tightly contacts the upper surface of the microplate 90A, thereby connecting the filter plate 30 and the microplate 90A in a liquid-tight state. The operations from dropping the sample liquid 80 into the first through-holes 312 to supplying the sample liquid 80 to the wells 91A by applying centrifugal force are the same as those in the first embodiment.

[0047] In the second embodiment, as in the first embodiment, a reinforcing sheet material 33 is disposed adjacent to the underside (the side from which the sample liquid flows out) of the filter sheet 32. Furthermore, the reinforcing sheet material 33 is not a mesh, but is a sheet material with multiple through-holes formed therein, and contacts the filter sheet 32 ​​by a surface rather than by a point or line. Therefore, during centrifugal filtration, the filter sheet 32 ​​is supported by the reinforcing sheet material 33 from the back side (the side facing the microplate 90A), thereby preventing damage to the filter sheet 32. Furthermore, because the reinforcing sheet material 33 has through-holes larger than the pore size of the first filter sheet, the sample liquid that has passed through the filter sheet 32 ​​is not prevented from being introduced into the wells 91A of the microplate 90A.

[0048] Furthermore, because the filter plate 30 is connected to the microplate 90A in a liquid-tight state by the flexible packing material 34, when the sample liquid 80 dropped into the first through-holes 312 is drawn into the microplate 90A by centrifugal force, the sample liquid is prevented from leaking and flowing into other wells 91A. Furthermore, because the cylinder 37 is provided on the underside of the base material 31 of the filter plate 30, the liquid sample that has passed through the filter sheet 32 ​​can be guided to near the bottom of the wells 91A of the microplate 90A.

[0049] In the filter plate 30 of the second embodiment, the filter sheet 32 ​​is sandwiched and fixed together with the reinforcing sheet material 33 between a first substrate 3111 and a second substrate 3112, and a filter is tightly stretched across each of the first through-holes 312. Therefore, in addition to filtering sample liquids, the filter plate 30 can be used to recover intracellular substances such as intracellular nucleic acids and exosomes contained in blood or cell culture fluid, for example.

[0050] When recovering intracellular substances from cells contained in a sample liquid, a filter sheet 32 ​​with a pore size of 1 μm or less is used, and the holder 701 of the centrifuge 70 is rotated at a higher speed than when filtering the sample liquid. This generates a large centrifugal force, forcing the cells in the sample liquid against the filter sheet 32. Because the filter sheet 32 ​​is tightly stretched across each first through-hole 312 of the filter plate 30, a large normal force acts on the cells pressed tightly against the filter sheet 32, and the centrifugal force and the normal force disrupt the cells. The disrupted cells and intracellular substances can be separated by appropriately setting the mesh size of the filter sheet 32. Alternatively, intracellular nucleic acids can be captured by using an appropriate filter sheet 32. In this way, when cells are pressed tightly against a filter sheet 32 ​​with a pore size of 1 μm or less while the filter sheet 32 ​​is tightly stretched, the filter sheet 32 ​​is particularly susceptible to damage. However, in the second embodiment, the filter sheet 32 ​​is supported by the reinforcing sheet material 33, so that damage to the filter can be prevented.

[0051] (3) Modifications The present invention is not limited to the above-described embodiment, and various modifications are possible.

[0052] The materials of the base material 11, 31, first filter sheet 12, 32, second filter sheet 13, reinforcing sheet material 33, and packing material 14, 34 shown in the above embodiments are merely examples, and other materials may be used. For example, ethylene propylene rubber (EPM (EPR) or EPDM (EPT)), urethane rubber, etc. may be used instead of silicone rubber for the packing material 14, 34. Furthermore, instead of nylon 66, the first filter sheet 12, 32 may be made of a polymer material such as polyester, polyethylene, or polypropylene, or may be made of a material other than a polymer material, such as metal. The PET sheet 131 of the second filter sheet 13 may be replaced by a sheet made of another material.

[0053] The mesh size of the first filter sheet 12, 32 can be adjusted to an appropriate size depending on the intended use of the filter plate 10, 30. For example, the mesh size can be 1 μm, 10 μm, 40 μm, 70 μm, 100 μm, etc. Alternatively, a microfiltration filter (membrane filter) with a mesh size of less than 1 μm (so-called submicron) may be used. For example, when recovering exosomes as intracellular substances, the mesh size of the filter sheet may be set to 0.1 μm to 5 μm. In either case, the second filter sheet 13 may have through-holes formed therein that are larger than the pore size of the first filter sheet 12, 32 but smaller than the diameter of the first through-holes 112, and the reinforcing sheet material 33 may also have through-holes formed therein that are larger than the pore size of the first filter sheet 12, 32 but smaller than the diameter of the first through-holes 312.

[0054] In the first embodiment, the second filter sheet 13 is used, and in the second embodiment, the reinforcing sheet material 33 is used as the support element of the present invention. However, various forms of support elements can be used. However, it is preferable to use a support element that contacts the first filter sheets 12 and 32 at a surface rather than at a point or line (i.e., supports the first filter sheets 12 and 32 at a surface). Furthermore, the second filter sheet 13 and the packing material 14 in the first embodiment, and the reinforcing sheet material 33 and the packing material 34 in the second embodiment, may be formed as a single component. In this case, the support element is made of a flexible material to function as a packing material. Furthermore, since it is expected that the through-holes will deform when the filter plate is pressed against the microplate, it is preferable to make the size of the through-holes formed in the support element larger than those described in the first and second embodiments to prevent this from interfering with the flow of sample liquid.

[0055] In the first and second embodiments, adhesive sheets that have been pre-attached are used as the second filter sheet 13 and the reinforcing sheet material 33, but the PET sheet 131 and the adhesive sheet may be prepared separately. Also, in the first and second embodiments, the filter and packing seal are integrally constructed using a double-sided adhesive film or an adhesive sheet, but these do not necessarily have to be integrated. As in the above embodiments, each component has a cutout, so by inserting each component into the microplate insertion space 115, 315 in order while aligning the cutout positions of the components, the components can be aligned without integrally constructing the filter and packing seal using a double-sided adhesive film.

[0056] In the first embodiment, a filter plate having first and third through-holes whose number and positions correspond to the wells 91 of a 96-well microplate 90 is shown, and in the second embodiment, a filter plate having first and third through-holes whose number and positions correspond to the wells 91A of a 384-well microplate 90A is shown. However, the filter plate 10 of the first embodiment and the filter plate 30 of the second embodiment can be applied to various microplates by providing first and third through-holes whose number and positions correspond to the wells 91, 91A of the microplates 90, 90A. Furthermore, the filter plate is not limited to a microplate, but can also be applied to dishes, tubes, bottles, flasks, bags, etc.

[0057] Furthermore, in the first and second embodiments, the first through-holes (substrate) and the third through-holes (gasket material) are provided at positions corresponding to all of the wells 91, 91A of the microplate 90, 90A, and the filter sheets 12, 13, 32 and the reinforcing sheet material 33 are arranged to cover all of those wells (sample storage sections). However, if only some of the wells (some of the multiple sample storage sections) are used, the first through-holes (substrate) and the third through-holes (gasket material) may be provided only at positions corresponding to those some of the wells, and the filter sheets 12, 13, 32 and the reinforcing sheet material 33 may be arranged to cover only the positions corresponding to those some of the wells.

[0058] In the filter plate of the first embodiment, corner portions 116 are provided in the microplate insertion space 115 of the substrate 11, but the corner portions 116 may be omitted. By omitting the corner portions 116, the filter plate can also be used for microplates that do not have a C-surface.

[0059] In the first and second embodiments, the filters corresponding to all the first through holes are constructed from a single first filter sheet 12, 32, and a single second filter sheet 13 and a reinforcing sheet material 33 are placed on the back surface thereof. However, as in the filter plate 10A shown in Figure 9, it is also possible to provide individual first filters 12A and second filters 13A (filters having a pore size larger than the pore size of the first filter 12A) for each of the multiple first through holes 112.

[0060] In the above embodiments, the planar shape of the first through holes 112, 312 is circular, but it may be quadrangular such as a square, hexagonal such as a regular hexagon, or other shapes. The diameters of the through holes described in the first and second embodiments are for circular through holes. In the case of through holes other than circular through holes, the through holes may have a size and shape that has the same area as the area of ​​a circle with the above diameter. Furthermore, the cross-sectional shape of the first through holes 112, 312 is not limited to the above examples.

[0061] (4) Related Embodiment Next, a filter plate 40 of a related embodiment will be described. The same components as those in the first embodiment will be denoted by the same reference numerals, and the description thereof will be omitted as appropriate.

[0062] Figure 10 shows a longitudinal cross-sectional view of a filter plate 40 according to a related embodiment. Figure 10 corresponds to the longitudinal cross-sectional view taken along line aa in Figure 1(a) described in the first embodiment. This filter plate 40 includes a substrate 11, a first filter sheet 42, a second filter sheet 43, and a packing material 14. The substrate 11 and the packing material 14 are the same as those in the first embodiment.

[0063] Fig. 11 is a vertical cross-sectional view showing a filter and packing seal 50 including a filter and packing material that is attached to a base material during the manufacture of the filter plate 40 of the related embodiment. Fig. 12 is an exploded perspective view of the filter and packing seal 50.

[0064] The first filter sheet 42 is the same material as the first filter sheet 12 in the first embodiment (although its arrangement differs from that in the first embodiment). The second filter sheet 43 is a sheet material in which an adhesive sheet 432 is bonded to one surface of a PET (polyethylene terephthalate) sheet 431 (one adhesive surface of a double-sided adhesive sheet is bonded to the PET sheet 431) and multiple through-holes are formed by irradiating the sheet with laser light. However, unlike the first embodiment, as shown in the partially enlarged vertical cross-sectional view of the second filter sheet 43 in FIG. 13 , the adhesive sheet 432 is located below the PET sheet 431. Furthermore, tapered through-holes 433 are formed, the diameter of which decreases from the top to the bottom (the adhesive sheet 432 side).

[0065] As shown in FIGS. 11 and 12 , the first filter sheet 42 and the packing material 14 are bonded to each other by a first double-sided adhesive film 45 having holes formed at positions corresponding to the third through holes 142 of the packing material 14 (the first double-sided adhesive film 45 is not shown in FIG. 10 ). The first double-sided adhesive film 45 may be the same as the first double-sided adhesive film 15 in the first embodiment. The first filter sheet 42 and the second filter sheet 43 are bonded to each other by an adhesive sheet 432 located on the underside of the second filter sheet 43. The second filter sheet 43 and the base material 11 are bonded to each other by a second double-sided adhesive film 46. The second double-sided adhesive film 46 may also be the same as the first double-sided adhesive film 16 in the first embodiment. The first double-sided adhesive film 45 and the second double-sided adhesive film 46 have the same planar shape as the first filter sheet 42, the second filter sheet 43, and the packing material 14. That is, two notches 423, 433, 143, 453, 463 are provided for each of the first filter sheet 42, the second filter sheet 43, the packing material 14, the first double-sided adhesive film 45, and the second double-sided adhesive film 46 at locations corresponding to the two C-faces of the base material 11 (see Figure 3).

[0066] The method of using the filter plate 40 of the related embodiment will be described below.

[0067] First, a 96-hole microplate is prepared that has a shape corresponding to the microplate insertion space 115 of the filter plate 40. Then, the C-face of this microplate is aligned with the corner portion 116 of the filter plate 40, and the microplate is inserted from below into the microplate insertion space 115 of the filter plate 40. Next, the microplate and the filter plate 40 are pressed against each other. At this time, because the packing material 14 located between the microplate and the base material 11 is flexible, the microplate 90 and the filter plate 40 are pressed against each other, and are therefore connected liquid-tightly with no gaps.

[0068] Next, pipette tips 60 are attached to each of the multiple nozzles of the multichannel micropipette, and sample liquid is aspirated into them. Each pipette tip 60 is then inserted into a first through-hole 112 in the substrate 11, and the tip of each pipette tip 60 is pressed against the second filter sheet 43 to dispense the sample liquid 80. This allows the sample liquid 80 to be simultaneously dispensed into multiple first through-holes 112 corresponding to multiple wells 91 arranged in a row. FIG. 14 is a partially enlarged view showing the process of inserting a pipette tip 60 into one first through-hole 112 and pressing it against the second filter sheet 43 to extract the sample liquid 80. This pushes the sample liquid 80 toward the first filter sheet 42, allowing the sample liquid 80 to be filtered by the first filter sheet 42 in a short time without using a centrifuge 70. While a preferred example of efficiently dispensing sample liquid 80 using a multichannel pipette with multiple nozzles has been described here, a pipette with only one nozzle may also be used.

[0069] As described above, dispensing sample liquid 80 by pressing the tip of the tip directly against a thin filter sheet is likely to damage the filter sheet. In contrast, in a related embodiment, a second filter sheet 43 is placed on top of the first filter sheet 42, preventing the tip of the pipette tip 60 from directly contacting the first filter sheet 42. This prevents damage to the first filter sheet 42. Furthermore, the through-holes in the second filter sheet 43 are tapered, decreasing in diameter from the top (the side where the sample liquid 80 flows in) to the bottom (the side where the sample liquid 80 flows out). In other words, the flow path for the sample liquid 80 gradually narrows. Therefore, the sample liquid 80 ejected from the tip of the pipette tip 60 is pressed against the first filter sheet 42 with great force, thereby filtering the sample liquid without using a centrifuge. The thickness of the second filter sheet 43 can be appropriately determined depending on the strength of the first filter sheet 42, the diameter of the through-holes formed in the second filter sheet 43, and other factors.

[0070] Aspects It will be understood by those skilled in the art that the exemplary embodiments described above are examples of the following aspects.

[0071] (Item 1) One aspect of the present invention is a filter plate that is attached to a sample container having a sample holding section that is open at the top, and includes: a) a substrate having a first through-hole formed at a position corresponding to the sample holding section, through which sample liquid flows; b) a first filter provided in the first through-hole; and c) a support that is positioned adjacent to the side of the first filter from which the sample liquid flows out, and that has a second through-hole formed therein that is larger in pore size than the first filter.

[0072] The filter plate according to paragraph 1 is fixed to a sample container so that the position of the opening of the sample container's sample storage portion coincides with the position of the first through-hole in the substrate, and the support is located closer to the sample container than the filter. In the filter plate according to the present invention, a filter used for centrifugal filtration is provided in the first through-hole of the substrate when fixed to the sample container. The filter plate according to paragraph 1 further includes a support positioned adjacent to the side of the first filter from which the sample liquid flows out. Therefore, when performing operations that apply force to the filter, such as centrifugal filtration, the filter is supported from the back side (the side facing the sample container), preventing damage to the first filter. Furthermore, the support has second through-holes formed therein that are larger than the pore size of the first filter used in centrifugal filtration, so that the sample liquid that has passed through the filter is not hindered from being introduced into the wells of the microplate.

[0073] (Clause 2) The filter plate according to clause 2 is the filter plate according to clause 1, wherein the filter plate is used for a microplate having a plurality of wells, the first through-holes are provided in the base material at positions corresponding to each well of the microplate, and the first filter is a sheet-like filter arranged on one side of the base material so as to cover all of the first through-holes.

[0074] The filter plate according to paragraph 1 can be used in a microplate having multiple wells, as described in paragraph 2. In this case, as described in paragraph 2, the first filter can be easily attached by using a sheet-like filter placed on one surface of the base so as to cover all of the first through-holes.

[0075] (Clause 3) A filter plate according to clause 3 is the filter plate according to clause 1 or 2, wherein the diameter of the second through hole is smaller than the diameter of the first through hole.

[0076] In the filter plate according to the third aspect, the filter can be supported by the support body inside the first through hole, regardless of the position where the second through hole is provided in the support body.

[0077] (Item 4) The filter plate according to item 4 is the filter plate according to any one of items 1 to 3, wherein the support is a second filter formed by laminating an adhesive sheet to a sheet-like material and forming the second through hole.

[0078] In the filter plate according to the fourth aspect, the first filter and the second filter are adhered to each other by the adhesive sheet of the support, thereby preventing the sample liquid from flowing between them and causing leakage.

[0079] (Item 5) The filter plate according to item 5 is the filter plate according to any one of items 1 to 4, wherein the diameter of the second through holes is 0.1 μm or more and 1 mm or less.

[0080] If the second through-hole is too small, the sample liquid that has passed through the first filter may have difficulty passing through the second filter. Furthermore, if the second through-hole is too large, the contact area between the first filter and the second filter will be reduced, narrowing the area that can support the first filter. Taking these factors into consideration, it is preferable that the diameter of the second through-hole be within the range specified in paragraph 6. Note that this assumes that the through-hole has a circular cross-section. In the case of a through-hole that is not circular, the size and shape of the through-hole should be the same as the area of ​​a circle with the above-mentioned diameter.

[0081] (Item 6) The filter plate according to item 6 is the filter plate according to any one of items 1 to 5, further comprising: a flexible plate-shaped packing material that is arranged on the side of the support from which the sample liquid flows out, and that has third through-holes formed at positions corresponding to each of the plurality of wells.

[0082] In the filter plate according to the sixth aspect, the sample liquid flowing in from the first through-holes of the substrate can be introduced into the wells corresponding to the first through-holes without leakage.

[0083] DESCRIPTION OF SYMBOLS 10, 10A, 30, 40...Filter plate 11...Substrate 111...Plate-shaped member 112...First through-hole 113...C-surface 114...Surrounding wall 115...Microplate insertion space 116...Corner portion 12, 42...First filter sheet 12A...First filter 13, 43...Second filter sheet 131, 431...PET sheet 132, 432...Adhesive sheet 13A...Second filter 14...Packing material 141...Plate-shaped member 142...Third through-hole 15...First double-sided adhesive film 16...Second double-sided adhesive film 20, 50...Seal with filter and packing material 31...Substrate 3111...First substrate 3112...Second substrate 3114...Surrounding wall 312...First through-hole 3121...Through-hole in first substrate 3122...Through-hole of second base material 315...Microplate insertion space 32...Filter sheet 33...Reinforcing sheet material 34...Gasket material 342...Third through-hole 37...Cylindrical body 433...Through-hole of second filter sheet (second through-hole) 60...Pipette tip 70...Centrifuge 701...Holder 71...Rotating shaft 80...Sample liquid 90, 90A...Microplate 91, 91A...Well

Claims

1. A filter plate to be attached to a sample container having a sample holding section with an open top, comprising: a) a substrate having a first through-hole formed at a position corresponding to the sample holding section, through which sample liquid flows; b) a first filter provided in the first through-hole; and c) a support positioned adjacent to the first filter on the side from which the sample liquid flows out, and having a second through-hole formed therein, the second through-hole having a pore size larger than that of the first filter.

2. The filter plate according to claim 1, characterized in that the filter plate is used for a microplate having multiple wells, the first through-holes are provided in the base material at positions corresponding to each well of the microplate, and the first filter is a sheet-like filter placed on one surface of the base material so as to cover all of the first through-holes.

3. The filter plate according to claim 1, wherein the diameter of the second through-holes is smaller than the diameter of the first through-holes.

4. The filter plate according to claim 1, wherein the support is a second filter formed by laminating an adhesive sheet to a sheet-like material and forming the second through-holes.

5. The filter plate according to claim 1, wherein the diameter of the second through holes is between 0.1 μm and 1 mm.

6. The filter plate according to claim 1, characterized in that it comprises a flexible, plate-like packing material arranged on the side of the support from which the sample liquid flows out, and having third through-holes formed at positions corresponding to each of the plurality of wells.

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