JP2006010437A - Liquid leakage preventive tool for microchip, and microchip holder - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a microchip holder and a liquid leakage preventive tool for a microchip having excellent X-Y-directional and Z-directional (vertical-directional) positioning characteristics for the microchip such as a cell chip, capable of immobilizing easily the cell chip, capable of preventing easily a testing liquid from being leaked, and capable of setting a using amount of the testing liquid to the minimum. <P>SOLUTION: A chip storage part for positioning the microchip is formed in a holder base member, and the liquid leakage preventive tool is provided to fix the microchip attached to the chip storage part while pressing from its upper side. A seal part is preferably formed between a surface of the microchip and the liquid leakage preventive tool to put the testing liquid for use in an opening part of the liquid leakage preventive tool. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a device for preventing leakage of a test solution for a microchip used for cell level inspection / analysis and the like, and a microchip holder using the same.

In the examination and analysis at the cell level, a microchip such as a microwell array chip (hereinafter also simply referred to as a cell chip if necessary) or a microreactor is used.
For example, taking lymphocytes as an example, when a pathogen such as a virus enters a human body, B lymphocytes and T lymphocytes detect the pathogen, and first increase the number of lymphocytes specific to the pathogen. It tries to protect the body by making a lot of antibodies and killing cells infected with viruses.
When such lymphocytes are efficiently detected from blood and analyzed for immune function, tens of thousands to hundreds of thousands of minute holes having a diameter of about 10 microns and a depth of about 15 to 20 microns are arranged. A microwell array chip is used.
In order to analyze lymphocytes effective against pathogens by placing one lymphocyte in one hole of this microwell array chip and arranging hundreds of thousands of lymphocytes, the hundreds of thousands of holes are addressed with a scanner. It is necessary to be able to read individual lymphocytes with a capillary equipped with means for selectively sucking out lymphocytes called a spotter, for example.

Up to now, silicon or resin microchips with microwells aligned on a substrate have been used as cell chips. As a method for fixing and holding these cell chips, the present applicant has proposed a method of adhering to a glass slide. investigated.
However, this method has revealed the following technical problems.

First, when the cell chip is bonded and fixed to the slide glass, it is difficult to uniquely determine the position in the XY direction.
In the data processing after the data was read by the scanner due to the deviation of the position of the cell chip in the XY direction, it was necessary to correct the deviation of the position in the XY direction, which took time for analysis.
Next, it became clear that adjustment in the height direction (Z direction) was also difficult.
The depth of focus of the scanner is +/- 50 microns, but the variation in horizontal height is large due to adhesive fixation, and the variation in height in the scanning range of the cell chip cannot be suppressed to within 60 microns. It was difficult to accurately acquire data in the scan range. Furthermore, when the lymphocytes are taken out from the individual holes of the cell chip by the automatic cell acquisition device, it is necessary to bring the tip of the spotter of the automatic cell acquisition device to the position of the individual holes. Since the position in the vertical direction is shifted, it is necessary to adjust the tip position of the spotter according to the position of the hole, and the operation becomes complicated.
Therefore, although it was considered to fix to the slide glass by a method other than adhesion, the slide glass itself has a thickness variation of about 200 microns, and it is difficult to reduce this. It turned out that adjustment in the vertical direction (Z direction) was difficult.

In addition, when a cell chip is fixed on a slide glass and a test solution is applied to stimulate with an antigen and a cover glass is applied, the solution leaks from the side and flows out, causing the required volume and concentration of the test solution. Setting was also difficult.
The applicants of the present application have accomplished the present invention as a result of intensive studies to solve the above technical problems.
Japanese Patent Laid-Open No. 2002-156381 discloses a technique related to a sample carrier provided with a cover that covers watertightness, but not only is the levelness insufficient, but the watertight structure is complicated.

Japanese Patent Laid-Open No. 2002-156381

  The present invention provides a microchip liquid leakage prevention device and a microchip such as a cell chip, which can easily prevent the leakage of the test liquid and can set the usage amount of the test liquid to the minimum under the background art as described above. An object of the present invention is to provide a microchip holder that is excellent in positioning in the XY direction and the Z direction (vertical direction) and that can easily fix a cell chip.

  The liquid leakage prevention device for a microchip according to the present invention comprises a resin-made or rubber-made frame body by molding, and when the flat attachment surface is formed on the frame body and attached to the surface of the microchip, An opening is formed in the frame so that at least a part of the minute recess of the chip faces.

In the present invention, the term “microchip” refers not only to the microwell array chip for immunological examination / analysis of lymphocytes described above as an example, but also to blood samples such as leukocytes and erythrocytes, or individual specimens having a cell level size. A microwell array chip for aligning with the microwells, a microreactor having a test chamber, a reaction chamber and a flow path.
Therefore, when a flat mounting surface is formed on a liquid leakage prevention tool made of a frame and mounted on the surface of the microchip, an opening in which at least a part of a micro concave portion of the microchip faces is formed in the frame. The microchip has micro-recesses such as well array regions and reaction chambers on the surface, but this opening can be formed by forming an opening in the frame that matches the minimum range required for testing. The minimum test solution is added to the test so that the test can be evaluated.
Therefore, the frame (outer shape of the liquid leakage prevention tool) can be a rectangular shape, a ring shape, or any other shape, but is generally matched to the planar shape of the microchip.
In addition, the shape and size of the opening formed in the frame can be appropriately selected and designed according to the well array region of the microchip, the number of wells required for the test, and the reaction mode in the microreactor.

  It is necessary to form a seal part on the mounting surface of the liquid leakage prevention tool because the mounting surface of the liquid leakage prevention tool is mounted on the surface of the microchip so that the test liquid is put into the opening. However, sealing properties can be easily secured by using resin or rubber.

The present invention further relates to a microchip holder. The liquid according to claim 1, wherein a chip storage portion for positioning the microchip is formed on the holder base member, and the microchip mounted on the chip storage portion is pressed and fixed from above. A leak prevention tool is used.
In the present invention, the microchip holder refers to a carrier holder for fixing and holding a microchip.
Here, the carrier holder means a holder for mounting a test sample on a test / evaluation apparatus.
The chip storage portion for positioning the microchip is used to position the microchip in the X, Y, and Z directions when the microchip is mounted. The chip storage portion is formed by digging up the upper surface of the holder base member. Alternatively, the chip housing portion may be formed by forming a protruding wall for positioning at least the corner portion of the microchip on the holder base portion.
Further, in this microchip holder, the liquid leakage prevention tool functions as a microchip pressing tool (hereinafter referred to as a liquid leakage prevention tool or a chip pressing tool as required).
Then, the cell chip is scanned from the opening formed in the frame body, and the capillary operation is performed.
Therefore, when the microchip is made of resin such as injection molding and the holder base member and the liquid leakage prevention tool are made of resin by molding, the height variation of the microchip can be kept within 60 microns. If the leak preventer is harder than the microchip, the microchip may be flattened.
If the microchip is made of silicon, when it is pressed and fixed with a liquid leakage prevention tool, stress is partially concentrated in the direction perpendicular to the surface and easily cracked. A step surface (receiving portion) was provided on the outer periphery of the microchip, and stress concentration on the microchip was prevented by receiving a pressing force by the liquid leakage prevention tool.
The holder base member and the liquid leakage prevention tool can be molded integrally or separately.

  In addition, when using the test solution in the opening of the frame, a cover glass is placed so that the test solution evaporates and the microchip does not dry. When the projection for positioning the cover glass is formed on the tool, the cover glass can be easily attached and detached.

In the present invention, the mounting surface of the liquid leakage prevention tool can be adhered and fixed to the surface of the microchip.
In addition, the liquid leakage prevention tool can be made to have a minimum test liquid amount by optimizing the shape, size, and depth of the opening.
In the present invention, since the chip storage portion is formed on the holder base member and the microchip is mounted and pressed by the liquid leakage prevention tool from above, the microchip is simply mounted on the chip storage portion in the XY direction. And positioning in the Z direction (vertical direction).
In addition, there is also an effect of improving the flatness of the upper surface of the microchip when the liquid leakage prevention tool presses the microchip against the holder base member.
Such a microchip holder can be easily manufactured with resin, and high flatness can be easily obtained by molding.
Further, since it can be mass-produced at low cost by molding, it can be used disposable, and when it is remelted and remolded, it is heated and sterilized so that it can be recycled.
The microchip holder according to the present invention has a high positional accuracy in the horizontal direction and the vertical direction, the focus is not shifted when scanning, the signal can be measured with high accuracy, and the position of the capillary or the like is easy to determine when collecting cells, Useful for automating cell recovery.

FIG. 1 shows an example of a chip holder 10 according to the present invention.
The chip holder 10 includes a holder base member 20 and a liquid leakage prevention tool (chip presser) 30.
FIG. 2 shows a plan view (b), a front view (b), and a side view (c) of the holder base member 20, respectively. FIG. 3 shows a plan view (b), a front view (b), Side views (c) are shown respectively.
The holder base member 20 forms a chip storage portion 21 and stores a resin-made microchip 40 having a well array region 41 having hundreds of thousands of minute holes.
In the example shown in FIG. 1, the holder base member 20 has a rectangular shape assuming a conventional versatile slide glass.
In addition, the chip storage portion is formed by a positioning wall 22 that matches the outer shape of the microchip. In the example shown in FIG. 1, the shape of the positioning wall 22 matches the outer shape of the liquid leakage preventer 30. Yes.
In addition, the surface of the bottom 23 of the chip storage portion may be flush with the upper surface of the holder base portion, or the chip storage portion may be formed by digging up the holder base member.
Further, when a partial opening 27 as shown by an imaginary line is formed in the bottom 23 of the chip storage portion as an example, scanning with transmitted light is possible in the case of a resin microchip.

The resin-made microchip 40 is stored in the chip storage portion 21, and the liquid leakage prevention tool 30 is fitted to the positioning wall 22 from above to fix the microchip.
Since the resin microchip is difficult to break, it can be easily fixed by simply press-fitting a chip presser (liquid leakage prevention tool) on the microchip in the chip storage portion.
The chip presser (liquid leakage prevention tool) 30 is injection-molded using ABS resin, forms an opening 31, puts test liquid into the opening 31, and covers the cover glass to prevent evaporation. (The cover glass is not shown).
Further, lymphocytes and cell information of each microwell addressed to the resin microchip 40 are taken out through the opening.
FIG. 4 is a sectional view showing a state in which the resin microchip 40 is fixed.
The microchip 40 is housed in the chip housing portion of the holder base member 20 and fixed with the liquid leakage prevention tool 30.
The inclined wall 32 forming the opening 31 of the liquid leakage preventing tool 30 for fixing the microchip is set to about 30 to 45 °.
The test solution S is put into the opening 31 surrounded by the inclined wall 32. The edge of the opening of the liquid leakage prevention tool 30 that presses the surface of the microchip 40 is a seal portion 33. Prevents leakage.
Therefore, since the test liquid reservoir is sealed with the liquid leakage prevention tool, the amount of the test liquid can be set to the minimum.

Various types of microchips have been developed for lymphocytes, blood tests, and cell test studies. In addition to the resin-made examples (PMMA and PC) shown above, various types such as silicon, silicon rubber, and glass can be used. In the case of rubber, PDMS can be used.
If a resin microchip is used, even if the resin chip is somewhat warped or deformed, if a resin leakage prevention tool with a higher hardness is used, the flatness (levelness) will be Acts in the direction of improvement.
For example, in the example shown in FIG. 1, when a resin microchip having a size of about 20 mm × 20 mm and a thickness of about 1 mm is used, and the thickness t of the liquid leakage prevention tool is about 1 mm, the chip is stored. The depth of the part is set to about 2 mm so that the liquid leakage prevention tool can be accommodated.
As a result, the liquid leakage prevention tool is pressed and fixed so as to improve the flatness of the microchip, and the opening of the liquid leakage prevention tool becomes the test liquid reservoir, so that the liquid does not leak from the seal portion 33.
The material of the holder base member 20 and the liquid leakage prevention tool 30 is not particularly limited, but is suitable for production by molding using a resin.
In particular, it is suitable to use a hard resin (hard resin such as styrene, ABS, MMA, polyamide, polypropylene, polyethylene, fluororesin, cyclic polyolefin, polycarbonate, etc.) to hold the microchip and to ensure sealing performance.
In the example shown in FIG. 1, the range of variation in the planar height of the upper surface of the microchip could be suppressed to within about 20 microns.
Accordingly, the microchip 40 can be positioned in the X and Y directions by the positioning wall 22 forming the chip storage portion, and the level of the flatness of the holder base member 20 and the liquid leakage prevention tool 30 is changed to the slide glass Compared to this, it has improved dramatically, making it easier to read the scanner and adjust the focus of the capillary.

FIG. 5 shows another embodiment of the chip holder.
If the chip housed in the chip housing portion 21a formed on the holder base member 20a is a silicon microchip 40a, it is easily broken by stress in the direction perpendicular to the plane, so that the silicon chip is pressed when the chip presser is pressed. The receiving portion 26a is provided so that excessive stress is not applied to the housing.
For example, a positioning wall 22a that matches the outer shape of the chip is formed, and a chip presser fixing wall 24a that is larger than that is formed separately.
The holder base member 20a in this case is shown as a plan view (b), a front view (b), and a side view (c) in FIG. 6, and the liquid leakage prevention tool 30a is shown in a plan view (b) and a front view (b) ) And side view (c).
FIG. 8 shows a cross-sectional view of the silicon microchip 40a in the fitted state, and the chip housing portion of the holder base member 20a has two steps.
Separately from the positioning wall of the silicon microchip 40a, the liquid leakage prevention tool 30a is fitted (press-fitted) and fixed to the chip presser fixing wall 24a.
A seal portion 33a formed at the edge of the opening 31a of the liquid leakage prevention tool 30a comes into contact with the surface of the microchip 40a to seal the test solution.
In this case, as shown in FIG. 5, a receiving portion 26a is formed on the outer periphery of the chip storage portion 21a, and the height of the receiving portion is substantially the same as the upper surface of the silicon microchip 40a, thereby preventing liquid leakage. Since the surface of the silicon microchip 40a is pressed by the inclined wall 32a of the tool 30a, stress is not concentrated on the silicon microchip.
The size of the silicon microchip 40a is about 20 mm × 11 mm.

FIG. 9 shows an example other than press-fitting as a method for fixing the liquid leakage prevention tool. When the silicon microchip 40a is housed and pressed by the liquid leakage prevention tool 30b from above, the side portion of the liquid leakage prevention tool 30b is shown. This is an example in which a locking projection 34b is formed on the inner surface of the extrusion frame fixing wall 24b and a locking recess 25b is formed inside the extrusion frame fixing wall 24b.
FIG. 9B shows an enlarged view of the engaging portion. The a dimension of the locking recess 25b is set to be slightly larger than the b dimension of the locking protrusion 34b, and the protrusion 34b is inserted into the chip presser. The possibility of deformation due to interference with the fixture fixing wall 24b is prevented.
Also, as shown in FIG. 9A, when the engaging portion is set diagonally in the vicinity of the corner portion of the liquid leakage prevention device 30b, the removal portion is pulled out from the corner portion without the protruding portion 34b when the liquid leakage prevention device is removed. be able to.

Next, application development examples such as a liquid leakage prevention tool and a positioning wall will be described.
The example shown in FIG. 10 is an example in which a chip presser (liquid leakage prevention tool) portion 30c is formed integrally with the holder base member 20c.
In this case, the opening part 31c is formed in the part used as the chip presser 30c, and the seal part 33c is formed in the back side of the inclination wall 32c.
Accordingly, the microchip is mounted on the storage portion 21c from the lower side, and the microchip 40 is fixed by the chip fixing plate 50 from the lower side.
A locking recess 51 is formed in the chip fixing plate 50 and is engaged with a locking projection 34c on the chip storage portion side.
As the engaging structure, the locking projections may be formed on either side.
Note that the test liquid does not leak to the outside because the seal portion 33c on the liquid leakage prevention tool side contacts the seal surface 52 of the chip fixing plate.
Further, when an opening is formed in the chip fixing plate 50, transmission scanning is also possible.

The example shown in FIG. 11 is an example in which positioning walls 22d for positioning the microchip 40 on the holder base member 20d are formed only at the diagonal corners.
In this case, the microchip 40 is attached to the chip storage portion 21d, and the locking protrusion 34d formed on the liquid leakage prevention tool 30d is engaged with the locking recess 25d formed on the positioning wall from above to thereby leak the liquid. The microchip is pressed and sealed by the seal portion 33d of the opening 31d of the preventer 30d.

The example shown in FIG. 12 is a development example of a method for fixing the liquid leakage prevention tool to the holder base member.
In the example shown in FIG. 1 or FIG. 5, the liquid leakage prevention tool is fitted (press-fitted) into the positioning wall 22 or the presser fixing wall 24a, and the microchip is pressed and sealed. The method shown in FIG. This is an example in which a locking recess 25e is formed on the front surface portion of the holder base member 20e, and is locked by a locking projection 34e formed on the back surface of the liquid leakage prevention tool 30e.
In this case, it is also the same that the locking projection is formed on the holder base member side and the locking recess is formed on the liquid leakage prevention device side.
In addition, a seal portion 33e is formed in the liquid leakage prevention tool 30e.

In the example shown in FIG. 13, the chip storage portion 21 f is formed into a concave shape, and the seal portion of the liquid leakage preventing tool 30 f is also inserted into the concave portion.
The method of vertically engaging the locking protrusion 34f of the liquid leakage preventing tool and the locking recess 25f of the holder base member 20f is the same as that shown in FIG.

  The example shown in FIG. 14 is a development example of a method for engaging the liquid leakage preventing tool 30g and the holder base member 20g. The locking protrusion 34g of the liquid leakage preventing tool 30g is inserted along the locking recess 25g in the vertical direction. It is a case to wear.

  The example shown in FIG. 15 shows an example in which the shape of the microchip storage portion 21h and the liquid leakage prevention tool (chip presser) 30h may be circular (ring shape).

The example shown in FIG. 16 shows an example in which positioning protrusions 61, 62, 63, 64 of the cover glass 60 are formed on the liquid leakage prevention tool 30.
When the test solution is used inside the opening of the liquid leakage prevention device, the cover glass 60 is used to close the opening of the liquid leakage prevention device so that the test solution is not evaporated and the microchip is not dried. Although it is placed, the cover glass positioning protrusions are formed on the liquid leakage prevention tool, so that the cover glass can be easily attached and detached.
In this case, it is preferable to form a projection for positioning the cover glass so that an opening d for injecting the test solution is formed.
FIG. 17A shows a modified example of the shape of the opening of the liquid leakage preventing device. When a small amount of test solution or the number of cells is sufficient, a circular shape 31f and an irregular shape 31e may be used.
FIG. 17B shows a case where the liquid leakage prevention tool is a microreactor 40b.

The example of the microchip holder which concerns on this invention is shown. The example of a holder base member is shown. An example of a liquid leakage prevention tool is shown. The expanded sectional view of the attachment figure of a microchip is shown. The other example of a chip holder is shown. The other example of a holder base member is shown. The other example of a liquid leak prevention tool is shown. The expanded sectional view of a microchip mounting part is shown. An example of the engagement structure of the liquid leakage prevention tool is shown. The example which formed the liquid leak prevention tool integrally in the holder base member is shown. An example in which a microchip positioning wall is formed only in a corner portion is shown. An example in which the liquid leakage prevention tool and the holder base member are engaged from above and below is shown. An example in which the liquid leakage prevention tool is engaged in the vertical direction while the microchip housing portion is formed in a concave shape is shown. An example in which the locking protrusion is inserted and fixed along the locking recess as a method for engaging the liquid leakage prevention tool and the holder base member will be described. An example in which the shape of the liquid leakage prevention tool is circular is shown. The example which provided the cover glass positioning protrusion in the liquid leak prevention tool is shown. The application example of the opening part of a liquid leak prevention tool is shown.

Explanation of symbols

10 Chip holder 20, 20a, 20b, 20c, 20d Holder base member 20e, 20f, 20g, 20h Holder base member 21, 21a, 21b, 21c Chip storage part 21d, 21e, 21f, 21h Chip storage part 22, 22a, 22b , 22d Positioning walls 23, 23a Tip storage portion bottom 24a Tip retainer fixing walls 25b, 25d, 25e, 25f, 25g Locking recesses 26a, 26b Chip retainer receiving surface 27 Holder base member openings 30, 30a, 30b , 30c, 30d Liquid leakage prevention tool (chip presser)
30e, 30f, 30g, 30h Liquid leakage prevention tool (chip presser)
31, 31a, 31b, 31c, 31d Opening 32, 32a, 32b, 32c, 32d, 31e, 31f of liquid leakage prevention device (chip presser) Inclined wall 33, 33a, 33b, 33c, 33d, 33e Seal portion 34b , 34 c, 34 d, 34 e, 34 f, 34 g Chip retainer locking projection 40 Resin microchip 40 a Silicon microchip 50 Chip fixing plate 51 Chip fixing plate locking recess 52 Chip fixing plate sealing surface 60 Cover glass 61, 62, 63, 64 Cover glass positioning protrusion formed on the liquid leakage prevention tool

Claims (6)

  A liquid leakage prevention tool comprising a resin-made or rubber-made frame body formed by molding, and when a flat mounting surface is formed on the frame body and mounted on the surface of the microchip, at least a minute recess of the microchip An anti-leakage device for a microchip, characterized in that an opening partly facing is formed in a frame.   2. The microchip liquid leakage preventer according to claim 1, wherein a seal portion is formed on the microchip mounting surface of the liquid leak preventer.   3. The microchip liquid leakage prevention device according to claim 1, wherein a projection for positioning the cover glass is formed.   2. A liquid leakage prevention device according to claim 1, wherein a chip storage portion for positioning the microchip is formed on the holder base member, and the microchip mounted on the chip storage portion is pressed and fixed from above. Microchip holder to be used.   The microchip holder according to claim 4, wherein the holder base member and the liquid leakage prevention tool are made of resin by molding.   6. The microchip holder according to claim 5, wherein the liquid leakage prevention tool is harder than the microchip.

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