JPS636832B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention involves sandwiching a thinly sliced biological tissue piece (referred to as a tissue section herein) between a slide glass and a cover glass, covering the tissue section with a mounting medium, and removing both glasses. The present invention relates to a microscopic specimen encapsulation device for producing a microscopic specimen by combining the two.

Testing and diagnosis through microscopic observation is widely used in various fields, but it is also used in the medical field for pathological tissue specimens, cytodiagnosis, blood tests, and various research and experiments. There is.

To prepare biological tissues into microscopic specimens, thin sections of biological tissues are pasted onto glass slides, and dyed with various dyes to make cells and tissue structures easier to see. When the material dries, it loses its transparency and becomes impossible to examine, so a mounting medium (an adhesive with good transparency and a refractive index close to that of glass dissolved in xylol, benzol, etc. to give it appropriate fluidity) is used. The specimen is then covered with a cover glass, and a encapsulation process is performed in which the tissue section is sealed between the two glasses.

The conventional mounting process will be explained with reference to FIGS. 1 and 2. First, a tissue section 2 attached to a rectangular slide glass 1 is dehydrated and stained, and then the tissue section 2 is stained using a xyol bath at the end of the staining process. While the xylene adhering to the tissue section 2 is not drying, an appropriate amount of mounting medium 4 adhering to the tip of a glass rod 3 is dropped onto the tissue section 2 as shown in FIG. The mounting medium spreads due to its surface tension.

Next, hold both ends of the cover glass 5 between fingers 6 as shown by the chain lines in FIG. I'm going to drop it on top of it. The mounting medium spreads due to its own surface tension, but by this operation it is pushed by the cover glass 5 and spreads further to completely cover the tissue section 2, and the section 2 is placed between the slide glass 1, the cover glass 5, and the mounting medium 4. As the mounting medium 4 solidifies, the cover glass 5 is adhered to the slide glass 1 and cannot be separated, completing a microscopic specimen.

If the amount of mounting medium is insufficient, inject it from the edge of the coverslip, and remove any excess mounting medium from the coverslip with a cloth soaked in xylol. Wipe it off with a piece. If mounting fails, place the slide glass in a xylol bath with the cover glass attached and remove the cover glass.

In addition to this, the mounting medium 4 may be dropped onto the cover glass 5, and the cover glass may be manipulated to cover the tissue section and overlap the slide glass.

In any case, the thickness of the cover glass is 0.06~
It is extremely thin, about 0.6 mm, fragile, and easily soiled, so you must be careful when handling it with your fingertips.

Also, during this encapsulation process, special care must be taken to prevent air bubbles from getting under the cover glass. If air bubbles appear, gently push the cover glass from above with tweezers to gradually push the air bubbles to one side and push them out of the cover glass. If air bubbles remain, they will prevent the normal passage of light and make microscopic observation impossible.

Conventional coverslipping processing was carried out manually as described above, which often broke the cover glass and left air bubbles, the amount of mounting medium dropped was inconsistent, and the operation required skill and was difficult to perform. It wasn't. In particular, it is difficult even for experts to completely prevent air bubbles from entering, and if air bubbles are introduced, pressing the cover glass from above to expel the air bubbles may have an adverse effect on cells and tissues. In addition, since the solvent used in the mounting medium is xylene or benzene, which has a refractive index close to that of glass, the vapor produced by manual mounting gives off a foul odor and is harmful if inhaled in large quantities, making it an undesirable process.

The present invention eliminates the above-mentioned inconvenience by automatically carrying out such a mounting process, and it not only makes the amount of mounting medium and stacking of cover glasses constant, but also makes the finished product of the microscope specimen constant. , was invented with the aim of obtaining a microscope specimen enclosure device that can prevent air bubbles from entering. The device that automatically performs the encapsulation process is based on U.S. Patent No.
3930928, Japanese Patent Publication No. 52-43106, and Japanese Unexamined Patent Publication No. 53-144761, these methods are similar to the present invention described below, in which a cover glass is moved into a slide glass by rotating an indenter. Rather than sliding it up along the edge of the glass slide and stacking it on the slide glass, it was constructed using a different method.

As schematically shown in FIG. 3, the apparatus of the present invention drips an appropriate amount of mounting medium from a thin nozzle 7 onto a slide glass 1 on which a tissue section 2 is pasted, and then attaches one end to the edge of the slide glass 1. By placing the cover glass 5 on the slide glass 5, the other end of which is lowered and supported by the bent part 8a of the indenter 8, by rotating the indenter 8 toward the slide glass 1, as shown in FIGS. 4a, b, and c. The cover glass 5 is stacked on the slide glass 1 while gradually reducing the angle to the slide glass 1 to expel air. As shown in Figure 5, if the mounting medium is dropped from multiple nozzles 7 arranged horizontally at several places on the slide glass, it will spread and form a pool of mounting medium that is long and continuous horizontally and wide. If a cover glass 5 is placed on top of this as shown in FIG. 4, the cover glass 5 can be placed on the slide glass 1 while covering the tissue section 2 and without intervening air bubbles. The slide glass 1 is preferably tilted slightly toward the cover glass 5.

The apparatus of the present invention will be explained below based on the illustrated embodiments. This embodiment is an example of the configuration of an apparatus for stacking a cover glass on a slide glass as shown in FIG. 4, and the apparatus of the present invention is not necessarily limited to this embodiment.

FIG. 6 shows an outline of the arrangement of each part of the apparatus, where A is the part where the cover glass 5 is overlapped with the slide glass 1;
B is a part that carries the cover glass to the part A, and C is a part that drops the mounting medium onto the slide glass. Each part is configured to operate in relation to each other and is mounted on the substrate 9. It will be done. Next, each part will be explained.

(1) Part A (part where the cover glass is overlapped with the slide glass) As shown in Figs. 7 to 9, the stand 10 on which the slide glass 1 is placed is a bracket 11 fixed to the substrate 9.
It is pivotally connected to the horizontal axis 12 by a horizontal shaft 12, and is formed into an inverted L-shape with a leg portion 10a that can rotate around the axis 12. The slide glass 1 is positioned in the width direction between four protrusions 10b and 10c provided on the top surface, and a stopper 1 is fixed to the side of the table.
3 to determine the position in the length direction and place it on the stand 10. The leg portion 10a has a base plate 9 and a spring 1.
4 and pulled to the right in FIG. 8, and after the straight-cut lower end surface of the base plate contacts the base plate 9, it becomes unrotatable and the base 10 stands upright in a fixed position. Also, due to the movement of the crank described later, the 8th
It is pushed down to the left in the figure. In the above-mentioned constant posture, the upper surface of the stand 10 is formed to be slightly inclined in the direction of the indenter 8.
The structure for placing the slide glass on the stand 10 in a constant position is the projection 10 described above.
It is not limited to b and 10c. In this embodiment, the operation of placing the slide glass 1 on the stand 10 is performed manually.

Four indenters 8 are connected by a shaft 15, and the narrow diameter portions at both ends of the shaft 15 are supported by a base 16. axis 1
A crank 17 is attached to one side protruding end of the base 16 of FIG. The pusher needle 8 is made to swing via the crank 17. One end of a push rod 21 is pivotally connected to the crank 17, and a spiral spring (not shown) provided at this pivot point exerts elasticity to rotate the push rod 21 counterclockwise in FIG. ing. As shown schematically in FIGS. 10 and 11, the middle part of the press rod 21 is supported by a support piece 22, and the tip of the press rod is pressed against a pin 23 planted on the side of the leg 10a of the stand 10. I have to. On the base 16, the lower end of the cover glass 5 that has been carried is placed on the eighth
As shown in FIG. 9, three needles 24 are installed between the pusher needles 8 in order to ensure that the pusher needles are positioned at the bent portion 8a and the upper side thereof is hung on the stand 10. There is.

When the crank 17 is rotated counterclockwise in FIG. 8 by the motor 18 and the connecting rod 20, and the pusher needle 8 is rotated counterclockwise, the cover glass 5 is moved onto the base 10 in the order shown in FIG. It is pushed up and stacked on top of the upper slide glass 1.

When the overlapping of the cover glass 5 and the slide glass 1 is completed and the pusher needle 8 is rotated clockwise in FIG. 8 to return to the initial cover glass receiving position, the push rod 21 moves to the tenth It is pushed out to the left as shown in the figure, and its end pushes the pin 23 to rotate the table 10 counterclockwise about the horizontal axis 12, and the slide glass on the table is moved to the pedestal 25.
drop it on top. Although the mounting medium has not yet solidified at this time, the cover glass is light so it will not move. Along with this movement, the height of the pin 23 decreases, so that after the slide glass has fallen, the push rod 21 is attached to the pin 23 as shown in Figure 11.
The leg 10a is pulled upward by the spring 14 and returned to its initial upright position.

(2) Part B (portion for transporting cover glasses) As shown in Figures 7 and 9, a large number of cover glasses 5 are stacked and stored in a fixed hopper 26, and a layer made of synthetic resin or the like is placed on top. A weight 27, which is not too heavy, is placed on it. The bottom of the hopper 26 supports the edge of the glass by bending the lower ends 26a of the front and rear side walls (left and right in FIG. 7) inward in a shelf-like manner in FIG. 9 to prevent the cover glass from falling. A gap is provided at the lower end of the side wall on the right side of FIG. 9 to allow only one lowermost cover glass to pass through to the right.

Below the hopper 26, a rolling element 29 to which a suction cup 28 is fixed is provided so as to move in the left and right directions in FIG. 9 (up and down in FIG. 7), first without rotating, and then while rotating. There is. That is, the rolling element 29
A gear 30 is connected to the gear 30, and the gear 30 meshes with a rack 31 located below. The rack 31 can slide in the left-right direction in FIG. 9 along the groove of the guide body 32 fixed to the base plate 9, and the
3, it is pushed towards the right edge of the groove. A stopper 3 is placed at the right end of the groove to prevent the rack 31 from coming out of the groove.
It is blocked by 4. The shaft of the gear 30 is the guide body 32
The motor 3 is supported by a bearing on a moving piece 35 that is movable in the left-right direction in FIG.
A gear 38 driven by gear 7 meshes with the rack 36. Therefore, when the motor 37 is rotated counterclockwise in FIG.
6 is sent to the right (downward in FIG. 7), and the movable piece 35, which is integral with the rack 36, moves to the right while bearing the gear 30. Since the gear 30 meshes with another rack 31, it rotates clockwise as the movable piece 35 moves to the right, and the rolling element 29 and suction cup 28 connected to the gear 30 also rotate at the same time. However, since it is configured as follows, the rolling elements 29 do not roll for the first time.

When the motor 37 is rotated in the opposite direction and the gear 38 is rotated in the opposite direction, the gear 30 and the rolling elements 29
returns to the left in FIG. 9 while rotating counterclockwise. Just before the rolling element 29 and gear 30 finish returning to the left, the contact piece 39 attached to the rolling element 29 hits the stopper 40 fixed to the moving piece 35, making the rolling element 29 and gear 30 unable to rotate. As the moving piece 35 moves to the left in FIG. 9 via the rack 36 due to the gear 38 continuing, the suction cup 28 and the rolling element 29 move to the left while compressing the spring 33 without rotating. Therefore, the right end of the rack 31 is slightly separated from the stopper 34.

Next, when moving the movable piece 35 to the right, the rack 31 must be moved until it hits the stopper 34.
1 does not rotate the gear 30, the suction cup 28 and the rolling element 29 move to the right without rotating until the rack 31 hits the stopper 34. Therefore, if the suction cup 28 is made parallel to the lowermost cover glass held by the hopper 26 at this time, the suction cup 28 will move when the rolling element 29 moves to the right.
The hopper 26 sucks the bottom cover glass 5 and moves it parallel to the other cover glasses.
It will be pulled out and then rotated.

The inside of the suction cup 28 is connected to a vacuum pump (not shown) through a flexible rubber tube 41, and a solenoid valve 42
Since the vacuum or atmospheric pressure is created by opening and closing the solenoid valve (Fig. 7), the lowermost cover glass in the hopper 26 is attracted to the suction cup 28 moving as described above and taken out by the operation of the solenoid valve. and is carried to the right in Figure 9. When the rolling element 29 advances to the rightward limit position, the suction cup 28 enters the state shown by the chain line in FIG. 9, brings the lower edge of the cover glass 5 close to the bent portion 8a of the pusher needle 8, and stops. This stop position and the return position where the suction cup stands upright under the hopper 26 are achieved by bringing the movable piece 35 into contact with the micro switches 43 and 44 to stop and rotate the motor 37 forward and backward.

When the suction cup 28 reaches the position indicated by the chain line in FIG.
Rubber tube 41 connected to the vacuum pump by switching
Atmospheric pressure is introduced into the chamber to release the suction of the suction cup 28. The cover glass 5 falls a little and is hung between the bent part 8a of the indenter and the stand 10. Thereafter, the cover glass 5 is successively overlapped with the slide glass 1 by the portion A mentioned above.

(3) Part C (part where mounting medium is dripped) As shown in FIG. It is branched into. The tube 45 communicates with a mounting medium tank 47 by a conduit in which there is a section of rubber tube 46 (FIG. 7).
As shown in FIG. 8, two pipes 48 and 49 are airtightly inserted into the mounting medium tank 47, which is airtightly covered. Compressed air is sent into the tank from the pipe 48 to drain the mounting medium liquid inside. Pressure is applied and the mounting medium is delivered from the tube 49 inserted close to the bottom. This pipe 49 passes through the rubber pipe 46 to the nozzle 7.
It communicates with a pipe 45 that connects the two.

A conduit 51 is provided in the conduit in front of the rubber tube 46 to open and close the flow of the mounting medium, and a gear 52 is connected to its shaft as shown in FIGS. 7 and 12. As shown in FIG. 12, it meshes with a gear 54 connected to a motor 53. The gear 52 further meshes with the gear 55,
The camshaft 56 supported by the base plate 9 is rotated in conjunction. A cam 57 (FIG. 7) is fixed to the camshaft 56 above the rubber tube 46, and the motor 53
After closing the pot 51 via the gears 54 and 52, the rubber tube 46 is compressed to force the mounting medium out of the nozzle 7 and dripped onto the slide glass 1, and then as the cam 57 rotates, the rubber tube 46 is compressed. When the rubber tube is released and restored by its own elasticity, the encapsulant that tends to drip from the nozzle 7 is sucked into the nozzle.

two other cams 58 coupled to the camshaft 56;
59 drives switches 60 and 61, respectively, to control the rotation of the motor 53 in a timely manner.
1, and the mounting medium is dripped from the nozzle.

At the rubber tube 46, there is a frame 64 with a U-shaped open leg end 62 pivotally connected to the substrate 9 by a horizontal shaft 63, and one end of the rubber tube 46 is fixed to the frame 64 by a holding piece 65. A rigid tube 4 with a branched mounting medium nozzle 7 is attached to the end of the rubber tube 46.
Connect the bent ends of step 5. A fork 66 (FIG. 12) is formed in the frame 64, and engages with a cam 67 provided on the camshaft 56. Therefore, the camshaft 56 rotates and the nozzle 7 is rotated as described above.
After dropping the mounting medium onto the slide glass 1, the tube 45 is lifted together with the tip of the frame 64 by the cam 67, so that the nozzle 7 gets in the way when the cover glass 5 is pushed up or the table 10 is rotated. Move it upward to prevent it from becoming a problem.

The above parts A to C are arranged in relation to each other on the substrate 9 as shown in FIG. 7, and a cover glass is attached. The operation order is as follows.

(1) Supplying compressed air to the mounting medium tank 47, filling the rubber tube 46, pipe 45, and nozzle 7 with the mounting medium, and starting the operation of the vacuum pump (not shown);
Preparations are made such as stacking cover glasses on the hopper 26.

(2) A glass slide 1, to which a tissue section 2 has been separately affixed and stained, is placed at a constant position on the table 10 in part A.

(3) Start the motor 53 and close the cock 51,
After making sure that the mounting medium in the rubber tube 46 does not retreat, the rubber tube 46 is pushed with the convex part of the cam 57, and the mounting medium is dripped from the nozzle 7 onto the tissue section 2 on the slide glass 1. When the convex part of the cam 57 separates from the rubber tube 46, the encapsulant at the end of the nozzle is sucked into the nozzle to prevent dripping, and then the cam 67 pushes up the frame 64 to raise the nozzle 7 a little. Avoid interfering with your work.

(4) Open the solenoid valve 42 to create a vacuum inside the suction cup 28,
The cover glass 5 at the bottom of the hopper 26 is adsorbed.

(5) Start the motor 37, drive the rack 36 with the gear 38, and move the rolling element 29 between the nozzle 7 and the platform 1.
Move in the direction of 0. As a result, stopper 3
The moving piece 35, which was a little far away from the hopper 4, advances until it hits the stopper 34. During this time, the rolling element 29 does not rotate and the cover glass 5 is pulled out from the hopper 26 in the horizontal direction, and the moving piece 35 reaches the stopper 34.
, the rolling element 29 starts rotating and tilts the suction cup 28 above the pusher needle 8 as shown by the chain lines in FIGS. becomes atmospheric pressure,
The cover glass 5 falls onto the indenter 8.

(6) When the movable piece 35 hits the micro switch 44 and the motor 37 rotates in reverse, and the suction cup 28 returns, the motor 18 starts rotating, the crank 17 is pulled by the connecting rod 20, and the pusher needle 8 rotates. The cover glass 5 is pushed up and stacked on top of the slide glass 1.

(7) In conjunction with the return movement of the pusher needle 8, the stand 10 is pushed by the push rod 21 and tilted, and the slide glass 1 with the cover glass 5 stacked thereon falls onto the pedestal 25.

(8) By repeating the above operations continuously or intermittently, a microscope specimen in which cover glasses 5 are stacked and bonded using a mounting medium is obtained. For intermittent operation, the motor 37 may be started manually.

As described above, the microscope specimen encapsulation device of the present invention automatically performs the encapsulation process, which was conventionally performed only manually, and can always perform encapsulation in a constant state, so that no air bubbles remain. There is almost no need to do anything, and good microscopic specimens can be obtained.

[Brief explanation of the drawing]

Fig. 1 is a schematic side view showing how mounting medium is dripped using a conventional method, Fig. 2 is a schematic side view showing a conventional manual operation in which a cover glass is placed on top of the mounting medium, and Fig. 3 and subsequent figures show examples of the present invention. 3 is a side view schematically showing how the cover glasses are stacked, FIG. 4 is a schematic side view showing the order of movement of the slide glass in this case, and FIG. 5 is a schematic plan view showing the state of mounting medium dripping. Figures 6 and below show the specific structure of the device. Figure 6 is a schematic plan view showing the arrangement of each part, Figure 7 is a plan view, Figure 8 is a right side view of Figure 7, and Figure 9. is an A-A view in FIG. 7, FIGS. 10 and 11 schematically show the mechanism for tilting the table 10, FIG. 10 is a schematic side view when it is upright, FIG. 11 is a schematic side view when it is tilted, and FIG. 12 is a schematic side view when it is tilted. FIG. 7 is a left side view of FIG. 7; 1: slide glass, 2: tissue section, 5: cover glass, 7: nozzle, 8: indenter, 8a: bent part, 10: stand, 15: shaft, 26: hopper.

Claims (1)

[Claims] 1. A table 10 is provided on which a slide glass 1 on which a tissue section is pasted and a mounting medium is dripped is placed, and a bent portion 8a is provided.
A shaft 15 parallel to the slide glass 1 on which a plurality of indenters 8, which are formed facing upward, protrude laterally, is connected to the base 1.
A cover glass 5, which is rotatably provided below the slide glass 1 on the stand 10 and the indenter 8 with the indenter side lowered, is gradually rotated by the rotation of the shaft 15. A microscope specimen encapsulating device characterized by being provided with a drive mechanism for sliding the specimen onto a slide glass 1 while reducing the inclination. 2. A table 10 is provided on which the slide glass 1 on which the tissue section is pasted and mounting medium is dripped is placed, and the bent portion 8a is
A shaft 15 parallel to the slide glass 1 on which a plurality of indenters 8, which are formed facing upward, protrude laterally, is connected to the base 1.
A cover glass 5, which is rotatably provided below the slide glass 1 on the stand 10 and the indenter 8 with the indenter side lowered, is gradually rotated by the rotation of the shaft 15. The cover glass 5 was taken out one by one from the hopper 26 into which the part A to be slid up on the slide glass 1 and the cover glass 5 were stacked while reducing the inclination, moved, and hung between the stand 10 and the pusher needle 8. The microscope specimen mounting device is composed of a part B for separating the cover glass 5 in the state, and a part C for sending a fixed amount of mounting medium to a nozzle 7 provided on a stand 10 and dropping it onto the slide glass 1.