JPH06323967A - Method and apparatus for obtaining and holding continuous thin chip for microscope - Google Patents

Abstract

PURPOSE:To make it possible to automate working and to save man power by synchronizing cutting with the movement of the sample of a microtome and the movement of tape, and sequentially lifting thin chips, which are thinly cut and floating on water surface with the planar state being kept. CONSTITUTION:Tape 5, which is supplied from a reel 4 in a tape box 3, passes through a tape guide 6 and dives into water 8 filled in a knife boat 7 for a microtome. The tape is lifted on the water before a knife 9. Meanwhile, a sample, which is attached to the microtome is cut with the lowering knife 9, and a thin chip 10 is obtained. The chips are expanded on the front surface of the water. Under the state, wherein the thin chip 10 is in contact with the tape 5, the tape 5 is lifted. Then, the thin chips 10 are mounted and aligned on the tape 5. The thin chips 10, which are cut with the knife 9, expand flatly on the water surface. At this time, the position of the tape guide 6 is changed back and forth, right and left and up and down with adjusting mechanisms 11, 12 and 13 so that the tape 5 comes to the tip of the thin chip 10.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention collects, moves or transfers a plurality of thin slices obtained by cutting an object in a plane with a knife to collect and hold thin slices for obtaining a continuous thin slice for a microscope. A method and a collecting and holding device.

[0002]

2. Description of the Related Art When a thin section sample of a microscope is made from a three-dimensional sample, a conventional wet microtome is used to make thin sections and collect them by cutting on a water surface stored in a knife boat in front of a knife. Squeezing the thin slices by hand using a delicate tool with a hair needle, or by grabbing the electron microscope mesh with tweezers and immersing it in water, gently pulling up and collecting the thin slices on it manually .

[0003]

Since the thin slices are very thin and brittle, it is difficult and time-consuming to obtain thin slices continuously by this conventional manual method. . Moreover, since the direction of the thin section is not determined, it was not suitable for producing a stereoscopic image from a microscopic image of the thin section.
Also, even if the cut thin individual slices are connected to form a ribbon by applying a paste such as paraffin to the upper surface of the sample block, it is possible to connect dozens of thin slices with one ribbon. there were. Moreover, the slicing work requires skill, and it was almost impossible to obtain several hundred or more continuous thin slices having a uniform thickness and aligned in direction.

Although recent advances in computer image processing techniques have made it easy to reconstruct the original three-dimensional morphology from a large number of consecutive slices, the above-mentioned conventional manual techniques described above: When it was moved or transferred to a microscope slide glass or an electron microscope mesh for observing it, it broke, shattered, turned upside down or out of order. Furthermore, variations in thin section size and the like occurred. Moreover, in order to avoid it, great care must be taken, and even making a few tens of continuous thin-section samples requires a great deal of time and labor. SUMMARY OF THE INVENTION In view of the above problems, it is an object of the present invention to provide a novel method for collecting and holding continuous thin slices for a microscope, which enables automation of work and labor saving, and an apparatus therefor.

[0005]

Therefore, in the method of the present invention, a microtome, which is an apparatus for making thin sections by cutting the surface of a three-dimensional sample at predetermined intervals, is used to supply water to a knife boat equipped with a fixed knife. In a so-called wet microtome that floats thin slices that have been collected and stored, prepare a long tape, guide this tape from above the water of the knife boat into the water, and further pull up the tape from this water. , Cutting from a three-dimensional sample by a microtome, cutting thin slices cut on the water surface of the knife boat with the tape, making one thin slice from the three-dimensional sample with the microtome, and thin slice By synchronizing the steps of scooping with tape alternately or simultaneously, it is possible to obtain a plurality of continuous thin samples from the three-dimensional sample. It is characterized in that to create a sequence of pieces on the tape.

Further, in the apparatus of the present invention, a microtome, which is an apparatus for cutting a surface of a three-dimensional sample at a predetermined interval to make a thin section, is a thin section cut by collecting water in a knife boat equipped with a fixed knife. In a so-called wet type microtome that floats, a means for supplying a long tape,
This tape is guided from the outside of the knife boat into the liquid of the knife boat and is configured to be folded back from the liquid at a position near the knife and pulled up, and a thin section is prepared from the three-dimensional sample with the knife by the microtome. The tape guide means is adapted to scoop up the thin section floating on the liquid surface with the tape.

[0007]

The operation according to the present invention is as follows. After the tape is introduced into the water of the water reservoir of the knife boat of the microtome, when it is pulled up onto the water, it absorbs the water and the surface of the water is raised. The cut thin section is pushed by the uncut sample block part during cutting to extend and the contracted thin section tries to extend flat on the water surface. The thin section is picked up on the tape while in contact with the tape, with a small amount of water between the tape and the thin section. By synchronizing the cutting with the movement of the sample of the microtome and the movement of the tape, the thin slices that are sliced one by one and float on the water surface are sequentially pulled up on the tape while maintaining the planar shape, and they are lined up like a frame of a photographic film It can be handled as a band lined up with. The thin section on the surface of the tape firmly adheres to the tape due to the slight evaporation of water in the gap between the thin section and the tape, and is not easily peeled off during the subsequent handling of the tape.

In order to arrange thin sections on a slide glass for microscopic observation, the slide glass coated with a thin section and the thin section attached to the tape are placed so that they face each other, and the gap is filled with water. If it is then dried, the thin slice is transferred to the glass, and even if the tape is removed, the thin slice does not come off from the glass.

[0009]

1 is a perspective view of an embodiment of the device of the present invention.
Shows an outline of a main part of the present invention. In FIG. 1, the device is composed of a screw 2 on the knife holder part 1 of the rotary microtome.
The tape 5 fixed by one and supplied from the reel 4 in the tape box 3 passes through several tape guides 6,
It is designed so that it can dive in the water 8 stored in the knife boat 7 for the microtome and be pulled up above the water before the knife 9. For that purpose, a folding back guide roller 6a for guiding the folding back of the tape is provided near the knife at the tip of the guide 6. When the sample attached to the microtome descends, the thin section 10 cut by the knife 9 spreads to the water surface in front of it, and when the thin section is in contact with the tape and the tape is pulled up, the thin section is arranged on the tape.

The thin section cut by the knife is flattened on the water surface after being cut, and at this time, the tape guide can be moved back and forth, left and right and up and down so that the tape comes to the tip of the thin section. , 12, 13 are provided. The pinch roller 14 for feeding the tape has a thin central portion so as not to touch the thin section on the tape. The rotation of the drive motor is transmitted through the flexible wire 23 to the gear 15
Rubber belt 1 to prevent further vibration
A tape-driven capstan roller 18 is rotated via 6 and a pulley 17.

FIG. 2 is a side view of the relationship between the tape and the sample. Again, as the sample 19 descends and is cut by the knife 9, the slice 10 spreads over the water surface of the boat and is then scooped up on tape.

FIG. 3 shows an example in which a photo interrupter 21 is used as an example of a mechanism for obtaining the timing for driving the tape. A shading plate 2 on the holder part of the sample 19
By attaching 0 and attaching the photo interrupter 21 to the microtome main body via the mechanism 21a capable of finely moving up and down, it is possible to obtain a tape drive timing suitable for the size of the sample and the slicing speed.

Reference numeral 22 in the drawing is a stepping motor,
Capstan roller 1 via flexible wire 23
Driving eight. FIG. 4 shows the positional relationship between the sample holder and the photo interrupter, and the knife and the tape. The advantage of using an optical sensor such as a photo interrupter is that the movement of the microtome is not loaded and the quality of the sliced piece is not affected. Although the sensor works even when the sample returns to the upper position, the motor control circuit 24 for driving the tape should be programmed not to drive the motor at this time, or the tape should be moved by half of the required amount by one response of the sensor. You can do it like this.

As a variation of the present invention, the tape material may be made transparent and the tape may be coated with an adhesive so that the tape is dyed. In addition, the mesh is arranged on a thin adhesive tape,
It is also possible to separate the samples one by one and use them as electron microscope samples by tape-forming them with a thin connection. At this time, it becomes a problem whether the mesh passes smoothly through the tape guide sunk at the front end of the boat, but when the mesh is attached to a thin adhesive tape to form a ribbon, it easily passes through a guide roller with a radius of 1.5 mm. Since it has been confirmed that the tape guide approaches at the tip of the boat, it only needs to be slightly deeper.

[0014]

The method and apparatus according to the present invention have the following effects. Without needing any special skill,
Sample preparation can be obtained quickly and easily. It is possible to make dozens or hundreds of continuous thin slices, which has been impossible so far, in a short time without loss. The device does not require microtome processing and can be easily installed on any model by using a clamp that matches the shape of the knife holder. It is possible to obtain thin slices with less distortion and inclination suitable for image processing. When this device is used for 1 to 2 micron thick slices of an optical microscope, the thin slices on the tape are not easily peeled off when the water attached when scooped up is dried, but with chrome gelatin etc. By arranging the tapes on the coated glass slides so that the tapes face each other, only the thin slices can be easily transferred. Unlike the thin slices that are manually arranged, the slope and arrangement are constant and suitable for computer image processing.

[Brief description of drawings]

FIG. 1 is a perspective view of a device of the present invention.

FIG. 2 is a side view showing the outline of the device of the present invention.

FIG. 3 is a perspective view of essential parts including a mechanism for obtaining a timing for driving a tape.

FIG. 4 shows a positional relationship between a sample holder and a photo interrupter, and a knife and a tape.

FIG. 5 is a view showing an electron microscope sample using a mesh.

FIG. 6 is a perspective view showing a state in which a thin section is inverted from a tape and attached to a slide glass.

[Explanation of symbols]

 1 Knife holder 2 Tape drive device fixing screw 3 Tape box 4 Tape reel 5 Tape 6 Tape guide 6a Folding guide roller 7 Knife boat 8 Water 9 Knife 10 Thin section 11 Left-right fine adjustment screw 12 Vertical adjustment screw 13 Vertical adjustment screw 14 Pinch roller 15 Gear 16 Rubber belt 17 Pulley 18 Capstan roller 19 Material block 20 Light shield attached to material holder 21 Photo interrupter 21a Fine movement manipulator 22 Stepping motor 23 Flexible wire 24 Motor control circuit

Claims (8)

[Claims] 1. A microtome, which is a device for making thin slices by cutting the surface of a three-dimensional sample at a predetermined interval, to float the cut thin slices by collecting water in a knife boat equipped with a fixed knife. In a wet microtome, prepare a long tape, guide this tape from above the water of the knife boat into water, and further pull up the tape from this water, cut from a three-dimensional sample with a microtome, A step of scooping a thin section cut on the water surface of the knife boat with the tape, a step of making one thin section from the three-dimensional sample with the microtome, and a step of scooping the thin section on the tape are alternately or simultaneously performed. Synchronizing creates a series of multiple thin slices on the tape from the three-dimensional sample. Settling holding method of a microscope for continuous thin sections characterized and. 2. The continuous thin slice for a microscope according to claim 1, wherein the tape and the thin slices arranged on the tape adhere to each other, and the tape serves as a support material when the thin slice is handled. Collection and retention method. 3. The sample of an electron microscope can be made by forming a shape and a material of the tape into a series of meshes on which a sample of an electron microscope is placed. Method for collecting and holding continuous thin sections for microscopes. 4. The transparent tape having a small optical strain is used as the long tape, and the thin sliced thin section can be observed under a microscope without being transferred to another tape. The method for collecting and holding continuous thin sections for a microscope according to claim 1. 5. A microtome, which is a device for making thin slices by cutting the surface of a three-dimensional sample at a predetermined interval, so as to float the cut thin slices by collecting water in a knife boat equipped with a fixed knife. In the wet microtome, a means for supplying a long tape, and the tape is configured to be guided from outside the knife boat into the liquid of the knife boat and folded back from the liquid near the knife and pulled up. A continuous guide thin film for a microscope, characterized in that a thin section is created from the three-dimensional sample by the microtome with the knife and the thin section floating on the liquid surface is picked up by the tape. Device for collecting and holding sections. 6. The microscope according to claim 5, further comprising means for making a series of a plurality of continuous thin slices from the three-dimensional sample on the tape by repeating the step of scooping the thin slices on a tape. Device for collecting and collecting continuous thin slices. 7. The apparatus for collecting and collecting continuous thin slices for a microscope according to claim 5, wherein the tape guide means is attachable to and detachable from a standardized knife of the microtome or a holder portion thereof. 8. A position detection sensor of the three-dimensional sample fixing base is mounted between a non-movable part of the main body of the microtome and a sample fixing base which makes periodic movements via a finely movable supporting base, and a signal thereof is transmitted. 6. The continuous thin section collecting and holding device for a microscope according to claim 5, wherein a cycle of sample cutting and a timing of scooping thin sections on a tape can be synchronized as a trigger.