JP3432173B2 - Biological tissue processing equipment - Google Patents

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a biological tissue processing apparatus, and more particularly to an immunostaining process and an ISH (in situ hybridy).
zation) The present invention relates to a treatment device for biological tissue.

[0002]

2. Description of the Related Art Conventionally, when performing biological tissue treatment such as immunostaining treatment or ISH treatment, a sample is attached to a slide glass, and the attached sample is subjected to many treatments such as dehydration, staining and hybridization. Is going.

Further, a DN produced on a slide glass
As a pretreatment for detection of the A chip, many reagent treatments such as denaturing with various reagents, hybridization, washing, and staining are performed.

Usually, in these treatments, about 20 slide glasses are arranged in a standing manner in a metal basket, and a plurality of glass water tanks containing various reagents are sequentially placed in order.
A series of treatments in which the basket is dipped is performed.
Then, the treatment of desired biological tissue or the treatment of the DNA chip is performed by the treatment with such various reagents.

Incidentally, it is known that irradiation with microwaves during an antigen-antibody reaction has the effect of increasing the frequency of collision between the two and increasing the speed of immune reaction. Furthermore, In S
Even in the case of in situ hybridization, irradiation with microwaves in the hybridization step
Similar effects are expected (reference: The Journal of His
tochemistry and Cytochemistry Vol.44, No.3, pp.281-2
87, 1996).

[0006]

The above-mentioned conventional processing apparatus,
That is, in the case of a device that performs processing using a metal basket that holds a plurality of slide glasses in an upright state, a water tank is used as described above, and the basket containing the slide glasses is immersed in the water tank. . Therefore, a large amount of reagent is required to immerse all slides in the reagent.

However, when immunostaining, ISH, etc. are performed, the reagents used for the treatment are very expensive, and in order to adapt to the economical conditions of the treatment,
It is necessary to treat with as little reagent as possible.
It is also desired from the environmental aspect to reduce the amount of waste liquid.

Further, it is necessary to set the holder in the water tank for each reagent, and the work becomes troublesome when the number of reagents to be processed increases. Further, in the case where a part of the sample is treated with a different type of reagent during a series of treatments, it is necessary to remove it from the holder, and there is a problem that work efficiency is extremely poor.

[0009] Further, in the research and examination in the field of gene diagnosis, more specimens are required to be prepared, and there is a problem in preparing a large number of specimens. Also, during the series of processing,
There is a step of treating with a very expensive reagent, and treatment with a smaller amount of reagent is desired.

Further, when performing microwave irradiation in order to shorten the processing time, it was necessary to take out the sample plate from the apparatus, perform appropriate processing and put it in a dedicated microwave irradiation apparatus. Note that related technology is disclosed in Japanese Patent Laid-Open No. 9-236598.

The present invention has been made to solve the above problems, and an object thereof is to treat a sample with a small amount of reagent and to easily exchange various reagents for each sample. It is to provide a processing device.

Another object of the present invention is to provide a biological tissue processing apparatus in which a sample plate can be easily handled.

Another object of the present invention is to provide an apparatus for treating biological tissue, which can treat with an extremely small amount of reagent such as a few drops.

Another object of the present invention is to provide a biological tissue processing apparatus having a function of automatically irradiating a microwave during a series of processing.

[0015]

To achieve the above object, the present invention provides a transparent sample plate to which a tissue sample is attached, a capsule into which the sample plate is dropped, and the capsule. A storage member having a plurality of storage portions for storing, wherein the tissue sample is treated with a reagent in the capsule , and the capsule is independent.
The capsule is configured as
And that the sample plate dropped into the inside can be transported.

According to the above construction, for example, a tissue sample in the form of a slice is attached to the surface of the sample plate, and the sample plate is dropped into the capsule. In this case, the sample plate is preferably stably held horizontally on the bottom surface of the capsule. Incidentally, it is desirable that the surface on which the tissue sample is attached face upward, but it may face downward. A reagent is dropped into the capsule, preferably by automatic dispensing, and a reagent process is executed. In this case, manual pipetting can be applied. After the treatment with the reagent, it is desirable to suction the residual reagent from the inside of the capsule and, if necessary, to wash the inside of the capsule. It should be noted that measures may be taken to protect the sample when a reagent is sucked.

Since the tissue sample can be handled while being carried on the sample plate, its handleability can be improved, and if necessary, the sample plate can be attached to a slide glass like a conventional cover glass, You can also create a slide. In this case, the attachment surface of the tissue sample is joined to the upper surface of the slide glass. Further, since the reagent processing can be performed in capsule units, the reagent processing can be realized for each tissue sample, and the amount of the reagent can be significantly reduced. Further, since the capsule itself is configured as an independent member, there is an advantage that the tissue sample can be transported together with the capsule. As a result, for example, the capsules can be transported to the microwave irradiation device.

As described above, the processing efficiency and operability can be improved by combining the sample plate and the capsule.

Desirably, the above-mentioned conveying means is a balloon which expands in the capsule and holds the capsule,
A drive unit that controls expansion and contraction of the balloon, and a moving mechanism that moves the balloon are included.

The use of such a balloon has an advantage that reliable transport can be performed without forming a special structure on the capsule. That is, since the capsule can be held simply by inflating the balloon in the space inside the capsule, no special engagement or grip is required. Even when capsules of various diameters are used, there is an advantage that they can be dealt with only by changing the size (pressure) of the balloon. Furthermore,
Even if a capsule having a non-circular cross section (for example, a rectangular shape) is used, the balloon itself can be deformed to reliably hold the capsule.

Preferably, the balloon is attached to a nozzle portion for dispensing, and inflation and deflation of the balloon is controlled by a dispensing pump connected to the nozzle portion. According to this configuration, the capsule transportation can be performed by effectively using the existing equipment, and the device cost can be reduced.

Preferably, a slit is formed in the capsule in the vertical direction. The slit functions as an opening for inserting tweezers, for example, when the sample plate is inserted into the capsule and when the sample plate is taken out from the capsule. When the reagent leaks to the outside through the slit, the reagent is retained in the container. A structure such as a reagent suction hole may be provided in the container.

Desirably, a cover member having a mesh for covering the sample plate is inserted into the capsule, and a reagent is dropped through the mesh. If the reagent is dropped onto the tissue sample through the mesh, there is an advantage that the amount of the reagent can be reduced by utilizing the development of the reagent on the mesh and a trace amount reagent can be supplied to the entire tissue sample.
In addition, the mesh has the advantage of protecting the tissue sample from reagent dripping.

Preferably, a cap having the same shape as that of the capsule is inserted into the capsule, and a gap for containing a tissue sample is formed between the upper surface of the sample plate and the lower surface of the cap. Especially when using highly volatile reagents, or when you want to prevent external contamination,
It is desirable to utilize the above cap. The shape of the cap may be a simple lid, but it is preferable to use a cap having an outer diameter slightly smaller than the inner diameter of the capsule and having the same shape as the capsule to be inserted into the capsule.

Desirably, the sample plate is bonded to a slide glass by using a surface on which a tissue sample is adhered as a bonding surface. That is, it is desirable to use the sample plate as a conventional cover glass. Therefore, the sample plate is formed as a material having a transparency and a thickness suitable for microscopic observation.

Preferably, the carrying mechanism carries the sample plate together with the capsule between the containing member and the microwave irradiation device.

[0027]

1 is a schematic configuration diagram of an apparatus according to the present invention. In FIG. 1, a plurality of accommodating portions 10A are formed in an array on a processing plate 10 as an accommodating member. Inside each accommodating portion 10A, a hollow capsule 11 having an opening formed in the upper portion is accommodated. Capsule 1
The sample plate 1 (see FIG. 2) is dropped on the bottom surface of the plate 1 and stably held horizontally. A sample 100 (see FIG. 2), which is a slice of biological tissue, is attached to each sample plate.

The dispensing nozzle 20 can be freely moved three-dimensionally by the transport mechanism 22. At the time of dispensing, the disposable tip 34A held by the tip rack 34 is attached to the tip of the dispensing nozzle 20. Then, one of the reagents 110A prepared in advance in the reagent table 110 is sucked by the chip 34A, and the reagent is dropped into all or some of the capsules 11 that require treatment with the reagent. With this device, it is possible to easily dispense a different reagent for each capsule, change the amount of reagent, mix a plurality of reagents in a sample capsule, and the like.

Dispensing nozzle 20 during capsule transportation
Instead of the disposable tip 34A,
The balloon body 80 held by the rack 84 is mounted.
The balloon body 80 functions as a capsule holding means as described later. The balloon body 80 is driven by the air pressure from the dispensing pump connected to the dispensing nozzle, and the balloon body is inflated / expanded by the electric control of the pump.
It is possible to easily control the contraction. Capsule 11
If the balloon body 80 is inflated in the inside thereof, the balloon body 80 is inflated according to the shape of the inner wall of the capsule 11, and the capsule 11 can be reliably held. After such holding, for example, the capsule 1
1 is transferred onto the microwave tray 71. Then, after the setting of the capsule 11 into the tray hole 71A is completed, the balloon body 80 is contracted, and then the microwave tray 71 is housed in the microwave irradiation device 70, and the tissue sample in the capsule 11 is irradiated with microwaves. Is executed. After the microwave irradiation, the sample capsule 11 is transported to the sample plate 10 again using the balloon 80,
Subsequently, reagent processing is performed. Note that, similarly to the above, if necessary, the capsule 11 is transported to a thermostatic bath (not shown) and subjected to rapid temperature treatment and the like.

The rack 92 holds a plurality of mesh capsules 90. The action of the mesh capsule 90 will be described later with reference to FIG. 4, but it is also carried by the balloon body 80 and inserted into the capsule 11 if necessary.

The processing plate 10 is controlled to a suitable temperature by a temperature control device 50 such as a heater, and the reagent in the capsule 11 is kept at a suitable reaction temperature. In a long-term reaction, a lid (cap) described later with reference to FIG. 8 is used to suppress evaporation of the reagent from inside the capsule 11.
The stirring means 40 rotates the processing plate 10 at an appropriate speed to stir the injected reagent and accelerate the reaction of the reagent.

When the reagent processing is completed, the sample capsule 11
The reagent therein is discharged by the suction unit 30. Here, the suction unit 30 is composed of a plurality of suction nozzles, which are freely transported by the transport mechanism 32.

According to the apparatus shown in FIG. 1, the reagent treatment of biological tissue can be automated including microwave irradiation and the like, and efficient treatment can be realized. In particular, there is an advantage that capsules can be individually transported by utilizing existing equipment.

FIG. 2 shows the setting state of the sample. Here, (A) shows the thin glass-like sample plate 1 to which the sample 100 is attached, and (B) shows a cross-sectional view of the capsule 11 which accommodates it therein,
In (C), a housing portion 10A in which the capsules 11 are individually housed
The processing plate 10 is shown in which is formed.

As the sample plate 1, for example, a commercially available transparent circular cover glass is used, and its dimensions are 0.2 mm in thickness and 15 mm in diameter. Sample plate 1
The surface of the sample is subjected to a treatment such as silane coating, and the sample 100 as a tissue section is attached.

The capsule 11 is a chemical-resistant resin molded product and has a cylindrical shape with an opening on the upper surface. The dimensions are, for example, an inner diameter of 15.5 mmφ and a height of 15 mm. The sample plate 1 is placed on the bottom surface thereof. If the horizontal cross-sectional shape of the capsule 11 is made to correspond to the shape of the sample plate 1, the reagent can be efficiently dipped in the sample when the reagent is dropped, and as a result, the amount of reagent necessary for the treatment can be reduced.

The processing plate 10 functions as a holding member for the capsules 11, and various forms can be adopted if the capsules are stably installed. The processing plate 10
Is made of, for example, Delrin, and the dimensions thereof are approximately the same as the microtiter plate: length 85 * width 125 * height 15 mm. The processing plate 10 has, for example, a diameter of 17 mm and a depth of 12 for mounting the sample capsule 11.
The cylindrical accommodating portion 10A having a size of 3 mm has 15 columns of 3 rows vertically * 5 columns
It is provided individually.

FIG. 3 shows a state in which the slit 11A is provided on the side surface of the capsule 11. The slit 11A is formed from the upper portion of the side surface to have a width of 3 mm and a height of 4 mm from the bottom surface, for example. Therefore, the processing plate 10
Slightly tilt the capsule 11 taken out from the slit 11
By inserting tweezers or the like into the capsule 11 via A, the sample plate can be easily taken out. The depth of the slit 11A may be set so that the reagent does not leak from the slit 11A, or
If the container 10A is positioned as the reagent processing container, the slit 1
1A may be formed up to the bottom surface of the capsule. Further, a hole may be formed in the bottom surface so that the sample plate can be pushed up from the lower side.

FIG. 4 shows the treatment of a tissue section with a very small amount of reagent by the capsule with mesh 90. The mesh capsule 90 is a cylindrical member having a bottom surface to which the mesh 90A is attached, and the dimensions thereof are, for example,
The outer shape is 14.5 mmφ and the height is 12 mm. As the mesh 90A attached to the bottom surface, for example, a mesh made of nylon and having openings of stitches of 40 μm is used.
Furthermore, in order to spread the reagent accurately, it is desirable that a hydrophilic treatment is applied. As shown in (A), when the capsule 90 with mesh is dropped into the capsule 11,
As shown in (B), the sample 100 is covered with the mesh 90A. In that state, for example, 20 μl
When a certain amount of reagent is dropped from above, the reagent is mesh 90
It penetrates into the space of the A stitch and spreads evenly on the tissue. Therefore, it is desirable to use the capsule with mesh 90 especially when using an expensive reagent which is sufficient for a small amount of treatment.

FIG. 5 shows an outline of the capsule holding method. The balloon body 80 includes a fitting portion 81 attached to the dispensing nozzle 20 and a balloon 82 attached to the fitting portion 81. By making the structure of the fitting part 81 the same as the upper part of the disposable chip, the balloon body 8 can be used by utilizing the existing chip attaching / detaching mechanism.
It is possible to attach and detach 0. As shown in (A), when gripping the capsule 11, air is supplied from the dispensing pump to the balloon 82 with the balloon 82 positioned in the center of the capsule. As shown in (B), the inflated balloon 82 securely holds the capsule 11. The removal operation can be easily performed by operating the dispensing pump in the suction direction and deflating the balloon 82.

FIG. 6 shows how the sample plate 1 is attached to the slide glass 102. An appropriate amount of a filler (eg, glycerin) is attached as the reagent 104 to the slide glass 102 in advance. The processed sample plate 1 is gently placed on the filler with the surface on which the sample 100 is attached facing downward so that no bubbles remain.
Slide glass 102 to which the sample plate 1 is attached
Is observed with an ordinary microscope as with ordinary glass slides.

FIG. 7 shows a cap 106 that functions as a lid for the capsule 11. The cap 106 is
A cylindrical main body 106 housed in the capsule 11 (inside the capsule 90 with mesh, if used).
C and the collar portion 1 that horizontally protrudes from the upper edge of the main body 106C.
06A and the bottom portion 106B of the main body 106C. An O-ring 108 is provided on the upper surface of the processing plate 10 so as to surround the upper opening of the accommodating portion, and the collar portion 106A is mounted on the O-ring 108,
The cap 106 is held. The bottom surface position of the bottom portion 106B is higher than that of the sample 100, and is slightly higher than that when the mesh 90A is used. That is, the bottom 10
A gap for containing the sample is formed between the lower surface of 6B and the upper surface of the sample plate 1.

The O-ring 108 is for enhancing the airtightness, and the O-ring may be attached to the collar portion 106A. Further, the collar portion 106A may be directly supported by the upper surface of the processing plate 10.

[0044]

As described above, according to the present invention,
There are advantages that the processing efficiency of a sample of biological tissue can be increased and the amount of reagents can be reduced.

[Brief description of drawings]

FIG. 1 is a conceptual diagram showing a biological tissue processing apparatus according to an embodiment.

FIG. 2 is a diagram showing a state when setting a capsule on a processing plate.

FIG. 3 is a conceptual diagram of a capsule in which a slit is formed.

FIG. 4 is a diagram showing a usage example of a capsule with mesh.

FIG. 5 is a diagram showing a method of transporting capsules using a balloon.

FIG. 6 is a diagram showing joining of a sample plate to a slide glass.

FIG. 7 is a diagram showing an example of use of a cap.

[Explanation of symbols]

1 sample plate, 10 treatment plate, 11 capsules, 20 dispensing nozzle, 70 microwave irradiation device, 8
0 balloon body, 90 mesh capsule, 110
Reagent table.

Continuation of the front page (56) References JP-A-3-85455 (JP, A) JP-A-8-327513 (JP, A) JP-A-9-281017 (JP, A) JP-A-7-195290 (JP , A) JP-A-8-40554 (JP, A) JP-A-8-285744 (JP, A) JP-A-9-236598 (JP, A) JP-A-9-43118 (JP, A) JP-A-9-43118 (JP, A) 3-176313 (JP, A) (58) Fields investigated (Int.Cl. ⁷ , DB name) G01N 1/28 C12M 1/00 C12M 1/42 G01N 1/30 G01N 33/48

Claims (9)

(57) [Claims] 1. A transparent sample plate to which a tissue sample is attached, a capsule into which the sample plate is dropped, and a storage member having a plurality of storage portions for storing the capsule. , The tissue sample is subjected to a reagent treatment in the capsule , and the capsule is configured as an independent member.
The whole capsule is dropped inside.
An apparatus for treating biological tissue, which is capable of transporting a sample plate . 2. The biological tissue processing apparatus according to claim 1, further comprising a transfer unit that transfers the sample plate for each capsule. 3. The apparatus according to claim 2, wherein the carrying means moves a balloon that expands in the capsule and holds the balloon, a drive unit that controls expansion and contraction of the balloon, and the balloon. And a moving mechanism for moving the biological tissue. 4. The apparatus according to claim 3, wherein the balloon is attached to a dispensing nozzle section, and inflation and deflation of the balloon are controlled by a dispensing pump connected to the nozzle section. Characterized biological tissue processing device. 5. The apparatus according to claim 1, wherein the capsule has a slit formed in a longitudinal direction. 6. The apparatus according to claim 1, wherein a cover member having a mesh that covers the sample plate is inserted into the capsule, and a reagent is dropped onto the tissue sample through the mesh. And biological tissue processing equipment. 7. The apparatus according to claim 1, wherein a cap having the same shape as that of the capsule is inserted into the capsule, and a gap for containing a tissue sample is provided between an upper surface of the sample plate and a lower surface of the cap. A device for treating biological tissue, which is formed. 8. The biological tissue processing apparatus according to claim 1, wherein the sample plate is bonded to a slide glass by using a tissue sample attachment surface as a bonding surface. The apparatus of claim 9 according to claim 2, wherein said transport mechanism processing apparatus biological tissue, characterized in that for conveying the capsules available and specimen plate between said housing member and the microwave irradiation apparatus.