JPH09275970A - Stirring apparatus for biochemical reaction - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a stirring apparatus for a biochemical reaction capable of effectively and uniformly stirring a very small amount of sample solution, etc., for an immune reaction, etc., without giving a physical damage to the sample. SOLUTION: This stirring apparatus for a biochemical reaction comprises a magnetic permeable vessel containing a magnetic substance in the interior thereof, a reacting vessel having arranged holes capable of supporting the magnetic permeable vessel and a magnetic device installed on the lower part of the reaction vessel and capable of acting a magnetic field on the magnetic substance. The magnetic device comprises magnet-retaining boards 7-9, supporting magnet pieces arranged in a circular shape corresponding to rows of the holes arranged in a concentric circle on the reacting vessel and comprising plural doughnut-like disks or one disk and at least one doughnut-like disk having a center of rotation the same as the reaction vessel and a driving means for rotationally driving the reaction vessel and/or the magnets-retaining boards 7-9.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a stirrer for a biochemical reaction, and to a stirrer capable of suitably stirring a solution that causes a biochemical reaction, or a biochemical reactor for performing a measurement operation including the stirring operation. It relates to a stirrer for a biochemical reactor used as part. More specifically, the present invention relates to antigen-antibody reaction, ligand-receptor reaction, DNA
The present invention relates to a stirring device for a biochemical reaction measuring device utilizing a biochemical reaction such as a hybridization reaction.

[0002]

2. Description of the Related Art Conventionally, for example, enzyme immunoassay for measuring a trace amount of biological components in a sample derived from blood has been widely and generally carried out in the clinical field. To describe one example, the immunoassay uses a labeled antigen or labeled antibody in which an appropriate label (fluorescent substance, radioactive substance, enzyme, etc.) is bound to either the antigen or the antibody, and an immune reaction in which the antigen and the antibody bind to each other. Then, a complex is formed with the component to be analyzed, and the amount of the component is measured by using the label in the complex as a guide. In the biochemical reaction represented by such an immunoassay, it is common to stir the components involved in the reaction for the purpose of appropriately causing the reaction to improve the measurement accuracy and shortening the measurement processing time. .

Further, in recent years, various biochemical reaction devices are arranged because a large number of reaction vessels are arranged on the circumference of a disc and the reaction vessel for transferring the reaction vessels by rotating the disc can be simplified in structure. Are frequently used by. In particular, a reaction vessel in which reaction vessels are concentrically arranged on a disc has the advantage that the radius of the disc does not increase even when a large number of vessels are held, and the size of the entire device can be made compact. ing.

[0004]

In a biochemical reaction such as an immune reaction, a sample to be subjected to the reaction is generally several ml.
It was difficult to use a device such as a stirring blade because the amount was as follows. Further, when the stirrer comes into contact with the sample (or the reaction solution containing the sample), other samples are contaminated due to carryover, which causes a wrong measurement result. Although a system in which the container itself is vibrated to perform stirring is also used, there is a problem such as scattering of the reaction solution despite the stirring efficiency. A method of stirring using ultrasonic waves has also been proposed, but in this case, components involved in the reaction may be physically damaged.

[0005]

The inventors of the present invention are suitable for a biochemical reaction apparatus having a disk-shaped reaction tank, and carry-over, scattering of reaction solution, physical obstruction of components involved in the reaction, etc. As a result of diligent examination of a stirring device that does not easily occur,
The present invention has been completed.

That is, according to the present invention, the magnetically permeable container, the magnetic body held inside the magnetically permeable container, and the holes capable of carrying the magnetically permeable container are (1) a disk-shaped structure in which a plurality of rows are concentrically arranged. A reaction tank, (2) a reaction tank comprising one disc having the holes arranged in one or more concentric circles and at least one donut-shaped disc, or (3) each concentric in one or more rows Two or more doughnut-shaped reaction tanks having the holes arranged therein, the reaction centers having the same center of rotation, and a magnetic body installed in the lower part of the reaction tank and held in the magnetically permeable container. A stirrer for biochemical reaction, comprising a magnet device capable of applying a magnetic field to the magnetic device, the magnet device carrying magnet pieces arranged in a circle corresponding to the row of holes arranged in the reaction tank. Multiple donors having the same center of rotation as the reaction tank A magnet holding plate made of a Jo or one disk and at least one donut-shaped disk, which is the apparatus characterized by being configured the reaction vessel and / or the magnet holding plate and a driving means for rotating movement. Hereinafter, the present invention will be described in detail. The magnetically permeable container holds a magnetic material described below, a sample solution to be stirred by a stirrer, and a sample solution containing a sample to be analyzed. Cup-shaped can be used. Also, an arcuate shape having a plurality of recesses can be used.

The one or more magnetic bodies held in the reaction vessel are not particularly limited as long as they can be moved by a magnet, such as iron and ferrite. Further, for example, the magnetic material may be bound with a resin binder or the like, the surface thereof may be coated with an appropriate material, or the magnetic material may be fused to the surface of the resin. As a suitable magnetic material, the magnetic beads described in Japanese Patent Publication No. 7-3425 can be exemplified. The shape of the magnetic material is particularly preferably a bead shape (spherical shape) that is easy to move in the reaction vessel. The particle size may be appropriately determined depending on the shape of the reaction vessel, the volume of the sample solution to be stirred, etc., but in the immunoassay in which a small amount of sample solution is stirred, it is usually 0.1 μm to 8 mm.
It is good to set it as a degree.

In a stirring device for an immunoassay device to which the present invention is preferably applied, components such as an antibody and an antigen, which are involved in an immune reaction, are preliminarily immobilized on a magnetic substance, or these components are mixed during a stirring reaction. The component may be immobilized. As described above, the magnetic substance of the present invention achieves stirring by moving in the reaction vessel, but at the same time, it can also have a function as a carrier for supporting the reaction components in the biochemical reaction.

The reaction tank is composed of (1) one disc, (2) one disc and at least one donut-shaped disc, or (3) two or more donut-shaped discs. In the case of (1), a plurality of rows of concentric holes are arranged on the surface, and in the cases of (2) and (3), one or more rows of concentric holes are arranged on the surface, respectively. This hole is for supporting the reaction vessel and does not need to penetrate the reaction vessel, but may be a recess formed on the reaction vessel and adapted to the bottom shape of the reaction vessel. When the holes do not penetrate the reaction layer, the reaction tank is made of a magnetically permeable member. Specifically, the reaction tank can be configured using a member such as plastic or aluminum.

In the present invention, the hole may penetrate the reaction tank, while the reaction vessel may have a shape that does not pass through the hole. For example, the reaction container may have a tapered tapered shape, or a protrusion may be provided on the reaction container to prevent the reaction container from dropping through the reaction layer. Further, for example, a thin plate for supporting the reaction container may be provided below the reaction layer. With this, it is possible to configure the reaction tank with various members without any particular limitation. Furthermore, the bottom of the reaction vessel and the magnet holding plate are not blocked, and the two are placed close to each other and the stirring efficiency by magnetic force is increased. Because it is possible to improve. In order to effectively act the magnetic action of the magnet pieces on the magnetic beads in the reaction vessel, it is preferable that the magnetic pieces passing under the reaction vessel and the magnetic beads be as close as possible. Depending on the magnetic force of the magnet piece, for example, 3 mm
It should be close to each other. The temperature of the reaction tank can be adjusted, and the temperature of the reaction solution in the reaction container can be controlled.

The number of holes and the number of rows thereof on the reaction vessel are determined so as to match the processing capacity required for the apparatus of the present invention, and if necessary, they may be arranged in multiple concentric circles on one disk, The doughnut-shaped reaction tanks may be arranged in one or a plurality of rows.

The magnet holding plate, which is installed in the lower part of the reaction vessel and has a magnetic field acting on the magnetic substance in the reaction vessel, is composed of a plurality of donut-shaped or one disc and at least one donut-shaped disc. The magnet pieces are arranged in a circular shape corresponding to the arranged row of holes, preferably at equal intervals in the circumferential direction. This magnet holding plate has the same center as the center of the reaction tank, and if one of the arranged magnet pieces is focused, the magnet piece is concentrically formed on the reaction tank by driving the magnet holding board, for example. It passes under a series of holes arranged.

It is possible to exemplify that the number of rows of the magnet pieces on the magnet holding plate is the same as the number of rows of the concentric holes provided in the reaction tank. With such a configuration, the reaction solutions in all the reaction vessels on the reaction tank can be stirred. Note that, for example, when the magnet pieces are preferably arranged in a staggered manner, it is apparent that a plurality of rows of magnet pieces are installed as the number of rows of holes in the reaction tank. On the other hand, for example, when stirring is performed only for a part of circular reaction solution in the reaction vessel held in concentric holes on the reaction tank, the row of holes in the reaction tank reaction vessel Fewer rows of magnet pieces are installed.

It is sufficient that the rows of the magnet pieces are arranged in a circle, for example, in a row at equal intervals in the circumferential direction, but in order to realize more efficient stirring, especially in a staggered manner. It is preferably arranged. By arranging the magnet pieces in a zigzag manner in this manner, the magnetic substance can be vibrated (reciprocating motion, circular motion, figure eight motion, etc.) at a predetermined frequency in the reaction container. The staggered arrangement may be appropriately determined based on the size of the bottom surface of the reaction container and the like. For example, when the bottom surface of the reaction container has a diameter of about 10 mm,
The zigzag arrangement of about 3 to 10 mm is preferable.

The installation interval of the magnet pieces may be determined so that the magnetic body moves (vibrates) at a predetermined frequency. The predetermined frequency is the volume of the sample solution to be stirred and the size of the magnetic body. It may be determined in consideration of the number of magnetic substances, but for example, in a normal immune reaction, 10 per minute
It can be exemplified that the number of times is about 300 times. An electric coil or the like can be used as the magnet piece itself,
Considering the problems of maintenance and heat generation, it is preferable to use a permanent magnet.

In the stirring apparatus of the present invention, the magnet holding plate is composed of two or more plates, each of which can be rotated at the same linear velocity. For example, it is possible to use a doughnut-shaped magnet holding board of the number corresponding to the number of rows of the magnet pieces, and carry one row of the magnet pieces on each of them and rotate them at the same linear velocity. In this case, even if the circumferential distance between the magnet pieces on each plate is the same, by rotating each plate at the same linear velocity, the hole array near the center point on the reaction tank and the center point The effect is that the degree of agitation with respect to the reaction container carried in the distant row of holes can be made uniform. Also, if you increase the linear velocity of a specific board,
It is also possible to increase the stirring efficiency with respect to the reaction container located in the row of holes on the reaction tank corresponding to the magnet pieces on the plate.

By rotating either or both of the reaction tank and the magnet holding plate to relatively rotate them,
The magnetic substance in the reaction container on the reaction tank can be moved. More specifically, the reaction tank may be rotated, the magnet holding plate may be rotated, or both may be rotated at different speeds in the same direction or different directions.

In the stirrer of the present invention, a magnet holding plate is located in the lower part of the reaction tank, and the upper space of the reaction tank is usually used for supplying the reaction container to the reaction tank and reagents necessary for biochemical reaction. Often used for addition. For this reason, the installation of the drive means for rotating the reaction tank is likely to be restricted and complicated. Therefore, it is preferable that the relative rotational movement is realized by moving the magnet holding plate. In this case, there is also an effect that operations such as addition of reagents to the reaction container can be made easier. For example, a rack gear may be provided at the bottom of each of the plurality of boards, and may be rotated by a set of a single or a plurality of pinion gears and a motor. By providing a gear having an equal pitch on each of the plurality of boards and rotating a pinion gear meshing with the gears and having an equal pitch and diameter by a motor or the like, each board can be rotationally driven at the same linear velocity.

[0019]

INDUSTRIAL APPLICABILITY The stirring device of the present invention is useful as a part of a biochemical reaction device such as an immune reaction measuring device, an enzyme reaction measuring device and an agglutination reaction measuring device, and also as an independent stirring device itself. . That is, according to the biochemical reaction stirring device of the present invention, the sample, the reaction solution, or the like can be effectively stirred by moving the magnetic substance in the reaction container carried in the hole on the reaction tank. In addition, by adjusting the relative rotational movement speed of the reaction tank and the magnet holding plate and the arrangement interval of the magnet pieces,
It is possible to easily change the degree of stirring.

Since the magnetic material used in the present invention is inexpensive, non-contact stirring for different reaction solutions can be performed by disposing each reaction container in a disposable manner, and carry-over between samples or reaction solutions can be prevented. . Further, in particular, if the structure is such that only the magnet holding plate is rotationally moved in a certain direction, the reaction container does not move, so that the reaction solution and the like in it does not scatter. Moreover, the degree of physical damage to the components involved in the biochemical reaction is extremely small as compared with ultrasonic stirring and the like, so that the present invention can be applied to stirring samples and reaction solutions containing various reaction components. Since the size of the magnetic substance can be selected according to the amount of the reaction solution or the like to be stirred, it is suitable for, for example, an immunoreaction measuring device or the like that needs to stir a trace amount solution of several ml or less. .

In the apparatus of the present invention, stirring can be performed by rotating the magnet holding plate installed in the lower part of the reaction vessel supporting the reaction vessel, so various operations for the reaction vessel, for example, dispensing of samples and reagents When carrying out operations, washing operations of the reaction container, and optical measurement operations for the reaction solution, it is possible to realize an arrangement that does not interfere with these operations. Moreover, if each magnet holding plate is configured to rotate at the same linear velocity, the row near the center of the reaction tank (inner side)
Achieves the same agitation regardless of the position where the reaction vessel is placed for the reaction solution etc. in the reaction vessel carried on the and the reaction solution etc. in the reaction vessel carried in the row farther from the center point (outer peripheral side) It is possible to

[0022]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in more detail with reference to the drawings, but the present invention is not limited to the invention described in these drawings.

FIG. 1 shows an example of an enzyme immunoassay apparatus constructed by using the stirring apparatus of the present invention, and is a view of the apparatus as viewed from above. The reaction vessel (1) has three concentric circular holes A provided on one disk-shaped reaction tank (2),
They are arranged like B and C. The reaction tank is kept at a constant temperature by a heater and a temperature sensor (not shown). As is clear from the figure, the triple concentric holes A, B and C
Each of the above is provided at a position where a transfer path of the reaction container by the reaction container transfer device (3) described later intersects with a circle on the reaction tank.

The reaction container (1) is a small container made of a black resin material and having a diameter of about 10 mm, a height of about 20 mm and an internal volume of about 2 ml. In the reaction vessel (1), a magnetic substance (plastic particles having a diameter of about 1.5 mm containing ferrite) on which an antibody to a substance to be analyzed is immobilized and an antibody to the component to be analyzed bound with an enzyme. Is charged, and a fixed amount of sample specimen (human serum sample) is dispensed by the liquid dispensing nozzle (17) prior to stirring. The specimen sample is dispensed at the sample dispensing position (D). The liquid dispensing nozzle 17 is connected to a syringe pump (not shown).

After dispensing the specimen sample, the reaction container (1) is transferred from the sample dispensing position D by the reaction container transfer device (3) into either row of B or C of the reaction tank which rotates at regular intervals. And held for a certain period of time. Then, the reaction vessel is transferred from the row B or C to row A by the reaction vessel transfer device (3). The reaction vessels supported in row A on this reaction tank are
By rotating the reaction tank (2) clockwise, a position (E) for washing and removing the unreacted enzyme-conjugated antibody-antigen complex, a position for dispensing the enzyme substrate (F) and remaining in the reaction vessel The enzyme activity in the immunocomplex is transferred to the position (G) where it is detected based on the color or fluorescence intensity of the reaction solution, and a predetermined operation is performed at each position for analysis.

In FIG. 1, 4 is the center of the reaction tank and a magnet holding plate (not shown), and 5 is the horizontal drive of the reaction vessel from the reaction vessel supply device (19) to the sample dispensing position D. A lane, 6 is a lane for driving the reaction container transfer device (3) in the horizontal direction, 18 is a sample supply device, and H is a position for discarding the used reaction container.

FIG. 2 shows a magnet holding plate (not shown in FIG. 1) and a driving means for rotating the magnet holding plate in a fixed direction. The magnet holding plate in this example is three donut-shaped plates corresponding to the three rows of holes on the reaction tank shown in FIG. 1, each carrying one row of magnet pieces. The center of each board (7-9) is the same as the center of the reaction tank, and they can rotate independently of each other. A large-diameter rack gear with teeth facing downward is provided under each board, and one pinion gear (10) having a shaft in the radial axis direction meshes with the gear of each donut board, and this pinion gear ( It rotates at the same linear velocity by the rotation of 10).

Here, if the diameter of the pinion gear is changed for each holding plate, it is possible to rotate them at different linear velocities. Although not particularly shown,
A large-diameter gear may be provided below the donut board so that the teeth are directed sideways, and a plurality of pinion gears each having a vertical axis may be engaged with each rack gear. in this case,
The plurality of pinion gears can be rotated by a single drive source, or can be rotated by different drive sources. If individual drive sources are used, it is possible to rotate each holding plate at an arbitrary linear velocity, or to stop some holding plates at an arbitrary time.

FIG. 3 shows a donut-shaped magnet holding plate (aluminum plate lined with a thin iron plate, 7 to 7) shown in FIG.
It is the figure which looked at 9) from the upper part (a) and the lower part (b). The magnet pieces (samarium-cobalt magnets) were pierced with φ5 × 3 holes in each donut disk, and were embedded in them to displace from the center circle (12) as shown in FIG. The distance to the center) was 3 mm in each of the inner and outer magnet pieces, and the magnet pieces were arranged in a zigzag pattern. This displacement was the same for all magnet holding plates.

In this example, the distance (7-1, 7-2, etc.) in the circumferential direction between the magnet pieces (11) arranged on the outer periphery of the donut-shaped disc 7 and the donut-shaped disc 7 are determined. The distance (7-3, 7-4, etc.) in the circumferential direction between the magnet pieces (13) arranged on the inner circumference is 18 mm and is the same. Also,
The circumferential distances (8-1, 8-2, 9-1, 9-2, etc.) between the magnet pieces in the other donut disks 8, 9 are also the same as 7-1, etc. in the donut disk 7.

FIG. 3b shows the gear (14) formed under each donut-shaped board, but the pitch of the gear crests of each donut-shaped board is the same. With such a configuration, due to the rotational movement of the donut plate, an equal oscillating magnetic field acts on the magnetic beads in the reaction container, and the magnetic beads move in the solution to achieve stirring. Since the magnets are arranged in a zigzag manner, the movement of the magnetic beads is a zigzag movement.

[Brief description of drawings]

FIG. 1 shows an example of an enzyme immunoassay device constructed using a stirring device of the present invention.

FIG. 2 shows a magnet holding plate and driving means for rotating the magnet holding plate in a fixed direction.

FIG. 3 is a diagram showing a donut disc-shaped magnet holding plate.

[Explanation of symbols]

DESCRIPTION OF SYMBOLS 1 reaction container 2 reaction tank 3 reaction container transfer device 4 center of a reaction tank and a magnet holding plate (not shown) 5 lane for horizontally transferring the reaction container 6 lane 7 for horizontally driving the reaction container transfer device , 8, 9 Donut disk-shaped magnet holding board 7-1 to 7-4, 8-1, 8-2, 9-1, 9-2 Distance in the circumferential direction of the magnet piece 7-5 Central circle of the magnet piece From 10 pinion gear 11 magnet piece (outer circumference side) 12 center circle 13 magnet piece (inner circumference side) 14 gear 17 sample dispensing nozzle 18 sample supply device 19 reaction container supply device A, B, C on reaction tank Rows of concentrically arranged holes D Sample dispensing position E Position for washing / removing unreacted enzyme-conjugated antibody-antigen complex F Dispensing position of enzyme substrate G Immune complex remaining in reaction container The enzyme activity in the body is determined by the color or fluorescence of the reaction sample. Position to detect based on strength H Position to dispose of used reaction vessel

Claims (10)

[Claims] 1. A magnetically permeable container, a magnetic body held inside the magnetically permeable container, and holes for carrying the magnetically permeable container (1) a disk-shaped reaction tank in which a plurality of rows are concentrically arranged. (2) A reaction tank comprising one disk having the holes arranged in one or more concentric circles and at least one donut-shaped disk, or (3) each being arranged in one or more concentric circles Two or more donut-shaped reaction vessels having the holes,
The biochemistry comprising a reaction tank having the same center of rotation and a magnet device installed at the bottom of the reaction tank and capable of exerting a magnetic field on the magnetic body held in the magnetically permeable container. A stirrer for reaction, wherein the magnet device carries a plurality of magnet pieces arranged in a circular shape corresponding to the row of holes arranged in the reaction tank, and has a plurality of donut shapes having the same center of rotation as the reaction tank. Alternatively, the apparatus comprises a magnet holding plate composed of one disk and at least one donut-shaped disk, and a driving means for rotating the reaction tank and / or the magnet holding disk. 2. The biochemical stirring device according to claim 1, wherein the magnetic substance is one or a plurality of beads on which a component that causes a biochemical reaction is immobilized or immobilized. 3. The stirrer for biochemical reaction according to claim 1, wherein the particle size of the magnetic material is 0.1 μm to 8 mm. 4. The biochemical reaction according to claim 1, wherein the magnet holding plate is concentrically arranged with the same number of magnet pieces as the number of rows of holes arranged concentrically in the reaction tank. Stirring device. 5. The raw material according to claim 1, wherein the magnet pieces arranged on the magnet holding plate are arranged in a circular shape and in a zigzag pattern corresponding to the row of holes arranged in the reaction tank. Stirrer for chemical reaction. 6. The stirring device for biochemical reaction according to claim 1, wherein the magnet pieces arranged on the magnet holding plate are arranged at equal intervals in the circumferential direction. 7. The stirrer for biochemical reaction according to claim 1, wherein the magnet is a permanent magnet. 8. The stirrer for biochemical reaction according to claim 1, wherein the driving means rotates the magnet holding plate at a constant speed. 9. The biochemical reaction stirring device according to claim 8, wherein the driving means comprises a rack gear provided on the magnet holding plate, a pinion gear meshing with the gear, and a means for rotating the pinion gear. 10. The biochemical reaction according to claim 8, wherein the driving means comprises a rack gear provided on each of the magnet holding plates, having a same pitch, a pinion gear meshing with the gear, and a means for rotating the pinion gear. Stirring device.