WO2025187033A1 - Filter plate - Google Patents

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

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

Traditionally, when a sample liquid, such as one derived from a living organism, is injected into the wells of a microplate, filters are used to remove impurities from the sample liquid and extract the target substance. The pore size of these filters is small, and the sample liquid often does not pass through the filter simply by dropping it onto the filter. For this reason, a process 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 (see, for example, Patent Documents 1 and 2).

Special Publication No. 2020-519440 Special Publication No. 2007-509633

When centrifugal filtering biological sample liquids, thin filters with small pore sizes are often used, which poses the problem of the filters being prone to tearing when subjected to operations that apply force, such as centrifugal filtration.

The problem that this invention aims to solve is to provide a filter plate that can prevent the filter from breaking when performing operations that apply force to the filter, such as centrifugal filtration of a sample liquid.

The present invention, which has been made to solve the above problems, provides a filter plate to be attached to a sample container having a sample storage portion with an open top, comprising:
a) a substrate having a first through-hole for passing a sample liquid at a position corresponding to the sample storage portion;
b) a first filter provided in the first through hole;
c) a support disposed adjacent to the first filter on the side from which the sample liquid flows out, and having second through-holes formed therein, the second through-holes having a pore size larger than that of the first filter.

The filter plate of the present invention is fixed to a sample container so that the position of the opening in the sample container's sample storage section coincides with the position of the first through-hole in the substrate, and so that the support is located closer to the sample container than the filter. In the filter plate of 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 of the present invention further includes a support positioned adjacent to the side of the first filter from which the sample liquid flows out. This allows the filter to be supported from the back side (the side facing the sample container) when performing operations that apply force to the filter, such as centrifugal filtration, thereby preventing damage to the first filter. Furthermore, the support is formed with a second through-hole that is 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 storage section of the sample container.

In the filter plate according to the present invention,
the filter plate is used in a microplate having a plurality of wells, and the first through-holes are provided in the base material at positions corresponding to the respective wells of the microplate;
The first filter is preferably a sheet-like filter disposed on one surface of the substrate so as to cover all of the first through-holes.

The filter plate of the present invention can be used in a microplate having multiple wells, as in the above embodiment. In that case, the first filter can be easily attached by using a sheet-like filter placed on one side of the base so as to cover all of the first through-holes, as in the above embodiment.

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 and forming the second through-holes therein.

In the filter plate of the above embodiment, the first filter and second filter are adhered together using the adhesive sheet of the support, preventing sample liquid from flowing between them.

The filter plate according to the present invention further comprises:
It is preferable that the support 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.

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

By using the filter plate of the present invention, damage to the filter can be reduced when performing operations that apply force to the filter, such as centrifugal filtration of sample liquid.

1A is a top view, FIG. 1B is a longitudinal cross-sectional view taken along line a-a, and FIG. 1C is a bottom view showing a first embodiment of a filter plate according to the present invention. FIG. 2 is a longitudinal cross-sectional view showing a filter/packing seal including a filter and a packing, which is attached to a base material during the manufacture of the filter plate of the first embodiment. FIG. 2 is an exploded perspective view of the filter/packing seal according to the first embodiment. FIG. 2 is a longitudinal cross-sectional view showing a state in which the filter plate of the first embodiment is attached to a microplate. FIG. 2 is a schematic diagram showing a state in which the microplate and the filter plate of the first embodiment are mounted in a centrifuge. 1A is a top view showing a second embodiment of a filter plate according to the present invention, FIG. 1B is a longitudinal cross-sectional view taken along line b-b, and FIG. 1C is a bottom view showing the second embodiment of a filter plate according to the present invention. 10A is a partially enlarged longitudinal cross-sectional view of a filter plate according to a second embodiment, and FIG. 10B is a partially enlarged bottom view of the filter plate. FIG. 10 is a partially enlarged longitudinal cross-sectional view showing a state in which the filter plate of the second embodiment is attached to a microplate. FIG. 10 is a partial vertical cross-sectional view showing a filter plate according to a modified example of the first embodiment. FIG. 2 is a longitudinal cross-sectional view of a filter plate according to an embodiment of the present invention. FIG. 10 is a longitudinal cross-sectional view showing a filter and packing seal including a filter and packing, which is attached to a base material during the manufacture of a filter plate according to a related embodiment. FIG. 10 is an exploded perspective view of a seal with filter packing material according to a related embodiment. FIG. 10 is a partial enlarged view of a second filter sheet in a related embodiment. FIG. 10 is a partially enlarged view showing how 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.

An embodiment of a filter plate according to the present invention will be described using Figures 1 to 9.

(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.

The base material 11 has a plastic plate-like member 111 that is rectangular in plan view, and a plurality of first through holes 112 that are circular in plan view and are provided in the plate-like member 111. A total of 96 first through holes 112 are arranged, with 12 arranged parallel to the long sides of the rectangle and 8 arranged parallel to the short sides. The arrangement of these 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 base material 11, the ends of the long sides are marked with numbers 1 through 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 through H corresponding to the rows in which the first through holes 112 are arranged. These numbers and letters are symbols used to identify each first through hole 112.

Of the four rectangular corners of the base material 11 (plate-shaped member 111), two adjacent corners across one of the short sides have C-shaped surfaces 113 cut off at a 45° angle relative to the short side. By providing the C-shaped surfaces 113, the orientation of the filter plate 10 can be easily identified.

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 arranged 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 arranged vertically as long as it surrounds the microplate insertion space 115.

The area below the bottom surface of the substrate 11 and surrounded by the surrounding wall 114 forms the microplate insertion space 115 into which a 96-well microplate is inserted. When a microplate is inserted 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 directly below the part of the rectangular parallelepiped where the substrate 11 is cut off at the C-face 113, a corner portion 116 (see Figure 1(c)) made of the same material as the surrounding wall 114 is provided. 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 approximately the same.

The first filter sheet 12 is made of a nylon 66 filter material with 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 base material 11 surrounded by the surrounding wall 114, and is attached to the underside of the base material 11 so as to cover that area. The portion of the first filter sheet 12 facing the first through-hole 112 functions as the filter in the present invention.

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 shown schematically only in Figure 3, and are not shown in Figures 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 base material 11, and is attached so as 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 formed 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 that are larger than the pore size of the first filter sheet 12 and smaller in diameter than the first through-holes 112.

If the through-holes in the second filter sheet 13 are too small, the sample liquid that has passed through the first filter sheet 12 may have difficulty passing through the second filter sheet 13. Furthermore, if the through-holes in the second filter sheet 13 are too large, the area of contact with the first filter sheet 12 will be reduced, resulting in a narrower area 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, regardless of the pore size of the first filter sheet 12, and more reliably supports the first filter sheet 12.

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 has a third through-hole 142 formed in a position corresponding to the first through-hole 112 of the base material 11. The packing material 14 has approximately 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.

2 and 3, the second filter sheet 13 and the packing material 14 are adhered 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 FIG. 1). Note that in FIG. 2, the horizontal and vertical scales are different (the vertical direction is longer) to clearly show each component. The first filter sheet 12 and the second filter sheet 13 are adhered 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 adhered 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 FIG. 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, and 163 are provided for each of the first filter sheet 12, second filter sheet 13, packing material 14, first double-sided adhesive film 15, and second double-sided adhesive film 16 at locations corresponding to the two C-faces of the base material 11 (see Figure 3).

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

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 and 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 and packing seal 20 and the manufacture of the filter plate 10, and then peeled off from the packing material 14 immediately before attaching the filter plate 10 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.

The following describes how to use the filter plate 10 of the first embodiment, with reference to Figures 4 and 5.

First, a 96-hole microplate 90 is prepared, shaped to fit the microplate insertion space 115 of the filter plate 10. Then, the C-face of this microplate 90 is aligned with the corner 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 (Figure 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 base material 11 is flexible, the microplate 90 and the filter plate 10 are pressed against each other, connecting them liquid-tightly and without any gaps.

Next, with the filter plate 10 fixed to the microplate 90, a micropipette (not shown) is used to drip a predetermined amount of sample liquid 80 into each of the first through-holes 112 in the substrate 11. At this time, it is sufficient that the dripped 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 drip sample liquid 80 into multiple first through-holes 112 simultaneously using a multi-channel micropipette with multiple nozzles.

After dropping the sample liquid 80 into the first through-holes 112 of the filter plate 10, the microplate 90 and filter plate 10 are tilted nearly vertically and set in the holder 701 of the centrifuge 70 (see Figure 5). To maintain close contact between the microplate 90 and filter plate 10 set in the holder 701 of the centrifuge 70, they may be fastened together with rubber bands, clips, etc. 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.

Filter plates come in a variety of pore sizes, and are used according to the intended purpose. For example, when centrifuging 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 like these are thin and weak, conventional filter plates can break when centrifugal filtration is performed.

In contrast, in the filter plate 10 of this embodiment, the second filter sheet 13 is placed adjacent to the underside (the side from which the sample liquid flows out) of the first filter sheet 12, which is used for the purpose of filtering the sample. Furthermore, the second filter sheet 13 is not mesh-like, but is made of sheet material with multiple through-holes formed therein, and is in contact with the first filter sheet 12 at a surface rather than at points or lines. Therefore, when centrifugal filtration is performed, the first filter sheet 12 is supported from the back side (the side facing the microplate 90) by the second filter sheet 13, preventing damage to the first filter sheet 12. Furthermore, because the second filter sheet 13 has through-holes formed that are 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.

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-hole 112 is drawn into a well 91 by centrifugal force, the sample liquid is prevented from flowing into other wells 91. This prevents different sample liquids from being mixed and supplied to the wells 91.

(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).

The filter plate 30 has a base material 31, a filter sheet 32, a reinforcing sheet material 33, a packing material 34, and a cylindrical body 37.

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

The substrate 31 consists of a first substrate 3111 and a second substrate 3112 attached to the bottom of the first substrate 3111. A rectangular frame-shaped surrounding wall 3114 extends downward from the outer periphery of the underside of the first substrate 3111. In this embodiment, the surrounding wall 3114 is also arranged 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 arranged vertically as long as it is arranged to surround the microplate insertion space 115.

The filter sheet 32, reinforcing sheet material 33, and second substrate 3112 are attached in this order from the first substrate 3111 side to the area surrounded by the surrounding wall 3114 on the underside of the first substrate 3111. The length of the surrounding wall 3114 is greater than the combined thickness of the filter sheet 32, reinforcing sheet material 33, and second substrate 3112, and the space below the second substrate 3112 and surrounded by the surrounding wall 3114 forms the microplate insertion space 315. The first substrate 3111 and the second substrate 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 substrate 3111 and the second substrate 3112 are stacked one on top of the other. Through-hole 3121 is cylindrical and has a uniform inner diameter in the depth direction (vertical direction in Figure 7(a)), whereas through-hole 3122 has a tapered shape with an inner diameter that is smaller at the bottom than at the top.

A cylinder 37 is provided on the underside of the second substrate 3112, extending downward from the periphery of the first through-hole 312. The cylinder 37 is made of the same plastic as the second substrate 3112 and is molded integrally with the second substrate 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 substrate 3112. The length of the cylinder 37 is shorter than the depth of the wells of a 384-well microplate.

The filter sheet 32 and reinforcing sheet material 33 are interposed between the first substrate 3111 and the second substrate 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 this 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, like the second filter sheet 13 in the first embodiment, a large number of through holes (corresponding to the second through holes in this 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.

Since 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, since 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 position at which the through holes are formed. In the second embodiment, for the reasons explained 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 the filter in the present invention.

The packing material 34 is made of a plate-shaped member made of silicone rubber and is attached to the second base material 3112 so as to cover the underside 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.

Although detailed explanations and illustrations are omitted, the first substrate 3111 and filter sheet 32, the reinforcing sheet material 33 and second substrate 3112, and the second substrate 3112 and packing material 34 are each bonded together using adhesive or an adhesive sheet with adhesive applied to both sides. The filter sheet 32 and reinforcing sheet material 33 are bonded together using an adhesive sheet provided on the reinforcing sheet material 33.

The filter plate 30 is used as follows. First, a microplate 90A (384 holes) having 384 wells 91A is prepared. Then, as shown in Figure 8, the microplate 90A is inserted from below into the microplate insertion space 315 of the filter plate 30, ensuring that the cylinder 37 corresponding to each well 91A of the microplate 90A is inserted. 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 adheres tightly to the top 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 in the first embodiment.

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 mesh-like, but is a sheet material with multiple through-holes formed therein, and is in contact with the filter sheet 32 by a surface rather than by points or lines. Therefore, when centrifugal filtration is performed, the filter sheet 32 is supported from the back side (the side facing the microplate 90A) by the reinforcing sheet material 33, preventing damage to the filter sheet 32. Furthermore, because the reinforcing sheet material 33 has through-holes formed that are 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.

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-hole 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 a 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.

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 the first substrate 3111 and the second substrate 3112, with the filter 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 fluids, for example.

When recovering intracellular material 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, pressing the cells in the sample liquid firmly 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 firmly against the filter sheet 32, and the centrifugal force and the normal force together destroy the cells. The destroyed cells and intracellular material 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 firmly 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, preventing damage to the filter.

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

The materials for 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, instead of silicone rubber, ethylene propylene rubber (EPM (EPR) or EPDM (EPT)), urethane rubber, etc. may be used for the packing material 14, 34. Furthermore, instead of nylon 66, the first filter sheet 12, 32 may be made of a polymeric material such as polyester, polyethylene, or polypropylene, or may be made of a material other than a polymeric material, such as metal. The PET sheet 131 of the second filter sheet 13 may be replaced by a sheet made of another material.

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) can be used. For example, when recovering exosomes as intracellular substances, the mesh size of the filter sheet should be 0.1 μm to 5 μm. In either case, the second filter sheet 13 should 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 should 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.

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

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 double-sided adhesive film or adhesive sheet, but these do not necessarily have to be integrated. As in the above embodiments, each component has a cutout, and so by inserting each component into the microplate insertion space 115, 315 in order while aligning the cutout positions of the components, it is possible to align the components without using double-sided adhesive film to integrally construct the filter and packing seal.

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.

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

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 with microplates that do not have a C-surface.

In the first and second embodiments, the filters corresponding to all the first through holes were constructed from a single first filter sheet 12, 32, with a single second filter sheet 13 and a reinforcing sheet material 33 placed on its back surface. 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 with pore sizes larger than the pore size of the first filter 12A) for each of the multiple first through holes 112.

In the above embodiment, the planar shape of the first through holes 112, 312 is circular, but it may also be quadrangular such as a square, hexagonal such as a regular hexagon, or any other shape. The diameter of the through holes described in the first and second embodiments is for when the through holes are circular; in the case of through holes other than circular, the through holes should be of 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.

(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.

Figure 10 shows a longitudinal cross-section of a filter plate 40 of a related embodiment. Figure 10 corresponds to the a-a longitudinal cross-section of Figure 1(a) described in the first embodiment. This filter plate 40 has a base material 11, a first filter sheet 42, a second filter sheet 43, and a packing material 14. The base material 11 and the packing material 14 are the same as those in the first embodiment.

Figure 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 a filter plate 40 of a related embodiment. Figure 12 is an exploded perspective view of the filter and packing seal 50.

The first filter sheet 42 is the same material as the first filter sheet 12 in the first embodiment (however, its arrangement differs from that of 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 Figure 13, the adhesive sheet 432 is located below the PET sheet 431. In addition, tapered through-holes 433 are formed, with their diameter decreasing from the top to the bottom (the adhesive sheet 432 side).

11 and 12, the first filter sheet 42 and the packing material 14 are adhered to each other by a first double-sided adhesive film 45 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 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 adhered 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 adhered to each other by a second double-sided adhesive film 46. The second double-sided adhesive film 46 may 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, and 463 are provided for each of the first filter sheet 42, second filter sheet 43, packing material 14, first double-sided adhesive film 45, and second double-sided adhesive film 46 at locations corresponding to the two C-faces of the base material 11 (see Figure 3).

The following describes how to use the filter plate 40 in the related embodiment.

First, a 96-hole microplate is prepared that is shaped to fit the microplate insertion space 115 of the filter plate 40. Then, the C-face of this microplate is aligned with the corner 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, connecting them in a liquid-tight state with no gaps.

Next, pipette tips 60 are attached to each of the multiple nozzles of the multi-channel micropipette, and sample liquid is aspirated into them. Each pipette tip 60 is then inserted into a first through-hole 112 in the base material 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. Figure 14 is a partially enlarged view showing how the pipette tip 60 is inserted into one first through-hole 112 and pressed 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 multi-channel pipette with multiple nozzles has been described here, a pipette with only one nozzle may also be used.

As described above, if the tip of the pipette tip 60 is pressed directly against a thin filter sheet to dispense the sample liquid 80, the filter sheet is likely to be damaged. 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 the first filter sheet 42 from being damaged. 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 set appropriately 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.

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

(Section 1)
One aspect of the present invention is a filter plate that is attached to a sample container having a sample storage portion with an open top, comprising:
a) a substrate having a first through-hole for passing a sample liquid at a position corresponding to the sample storage portion;
b) a first filter provided in the first through hole;
c) a support disposed adjacent to the first filter on the side from which the sample liquid flows out, and having second through-holes formed therein, the second through-holes having a pore size larger than that of the first filter.

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 section coincides with the position of the first through-hole in the substrate, and so that 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 in the substrate that is 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 is formed with second through-holes that are larger than the pore size of the first filter used for 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.

(Section 2)
The filter plate according to paragraph 2 is the filter plate according to paragraph 1,
the filter plate is used in a microplate having a plurality of wells, and the first through-holes are provided in the base material at positions corresponding to the respective wells of the microplate;
The first filter is a sheet-like filter disposed on one surface of the substrate so as to cover all of the first through-holes.

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

(Section 3)
The filter plate according to paragraph 3 is the filter plate according to paragraph 1 or 2,
The diameter of the second through hole is smaller than the diameter of the first through hole.

In the filter plate according to paragraph 3, the filter can be supported by the support body inside the first through hole, regardless of the position of the second through hole in the support body.

(Section 4)
The filter plate according to paragraph 4 is the filter plate according to any one of paragraphs 1 to 3,
The support is a second filter formed by laminating an adhesive sheet to a sheet-like material and forming the second through-holes therein.

In the filter plate according to paragraph 4, the first filter and second filter are adhered together using the adhesive sheet of the support, which prevents sample liquid from flowing between the two and causing leakage.

(Section 5)
A filter plate according to claim 5 is a filter plate according to any one of claims 1 to 4,
The diameter of the second through hole is not less than 0.1 μm and not more than 1 mm.

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 small, narrowing the area that can support the first filter. Taking these factors into consideration, it is preferable to set the diameter of the second through-hole 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 diameter.

(Section 6)
A filter plate according to a sixth aspect of the present invention is the filter plate according to any one of the first to fifth aspects, further comprising:
The support further comprises a flexible, plate-shaped packing material that is disposed on the side of the support from which the sample liquid flows out and has third through-holes formed at positions corresponding to the plurality of wells.

In the filter plate according to paragraph 6, the sample liquid flowing in from the first through-hole in the substrate can be introduced into the well corresponding to the first through-hole without leakage.

10, 10A, 30, 40... Filter plate 11... Base material 111... Plate-like 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-like member 142... Third through-hole 15... First Double-sided adhesive film 16... second double-sided adhesive film 20, 50... filter/packing seal 31... substrate 3111... first substrate 3112... second substrate 3114... surrounding wall 312... first through-hole 3121... through-hole 3122 of first substrate... through-hole 315 of second substrate... microplate insertion space 32... filter sheet 33... reinforcing sheet material 34... packing 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 solution 90, 90A... Microplate 91, 91A... Well

Claims (6)

A filter plate to be attached to a sample container having a sample receiving portion with an open top,
a) a substrate having a first through-hole for passing a sample liquid at a position corresponding to the sample storage portion;
b) a first filter provided in the first through hole;
c) a support disposed adjacent to the first filter on the side from which the sample liquid flows out, and having second through holes formed therein, the second through holes having a pore size larger than that of the first filter. the filter plate is used in a microplate having a plurality of wells, and the first through-holes are provided in the base material at positions corresponding to the respective wells of the microplate;
The filter plate according to claim 1, wherein the first filter is a sheet-like filter disposed on one surface of the base so as to cover all of the first through-holes. The diameter of the second through hole is smaller than the diameter of the first through hole.
2. The filter plate according to claim 1, characterized in that: 2. 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 therein. The filter plate according to claim 1 , wherein the diameter of the second through holes is 0.1 μm or more and 1 mm or less. 2. The filter plate according to claim 1, further comprising: a flexible, plate-shaped packing material disposed on the side of the support from which the sample liquid flows out, the packing material having third through holes at positions corresponding to each of the plurality of wells.