Ultra-thin microtome and its knife table

Abstract

The present invention belongs to the field of experimental instruments, and specifically provides an ultramicrotome and its knife table. The present invention aims to solve the problem that the section cut by the existing ultramicrotome has a short stroke and cannot meet the scientific research needs. To this end, the knife table of the present invention includes a knife table base fixedly connected to the base of the ultramicrotome, a knife table body slidably connected to the knife table base, a piezoelectric ceramic sheet provided on the knife table body, and a knife holder connected to the knife table body. Among them, when the piezoelectric ceramic sheet is energized, it can generate a deformation of 1 mm, and when the piezoelectric ceramic sheet is energized, it can drive the knife holder and the cutting tool mounted on the knife holder to move towards the sample arm and cut the sample on the sample arm. The section cut by the ultramicrotome with the above knife table of the present invention can reach 1 mm, meeting the needs of scientific research personnel.

Description

Technical Field

[0001] The present invention belongs to the field of experimental instruments, and particularly provides an ultramicrotome and a knife table thereof. Background Art

[0002] With the in-depth research of neurobiology and cell biology, especially the research related to the brain project proposed in recent years. In early 2016, the US Brain Project allocated 28 million US dollars to support Harvard University to obtain the structural and functional data of synaptic connections in 1 cubic millimeter of mouse brain cortex, and carry out research on the neuronal connections in the visual cortex for the research and development of a brain-inspired intelligent computing system. Researchers need to obtain three-dimensional structural information at the cell and even tissue level with a larger scale of cubic millimeter level, which relies on a new technology in electron microscopy - the method of serial section scanning electron microscopy imaging (ssSEM). This technology mainly automatically collects serial ultrathin sections of resin-embedded samples through a collecting belt, a diamond knife and an ultramicrotome, then significantly processes the sections by plasma thinning, measures them with an atomic force microscope, coats the surface of the sections with a conductive film and transfers them to a scanning electron microscope, and then performs high-resolution large-scale three-dimensional reconstruction on the obtained series of electron microscopy images. Its unique advantage is that serial sections can be preserved and can be imaged multiple times at different resolutions. Therefore, some precious samples can be repeatedly "consulted" by researchers like books in a library, and researchers can conduct sub-studies on the sections of interest, thus improving the acquisition efficiency and flexibility.

[0003] Among them, an ultramicrotome is a slicing machine used to make ultrathin sections for an electron microscope. It can cut samples embedded with various embedding agents into ultrathin sections less than 50 nanometers with a glass knife or a diamond knife. Ultramicrotomes mainly include two types: mechanical propulsion type and metal thermal expansion type.

[0004] The maximum single stroke of the existing ultramicrotome for preparing ultrathin sections (the thickness of the ultrathin sections is at the nanometer level) is limited by the stroke of the sample arm, usually 200 μm. When the slicing stroke reaches the upper limit, the sample arm automatically resets and returns to the starting position of the stroke. If ultrathin sections with a thickness exceeding 200 μm are desired, the knife must be realigned after the sample arm is reset. For the collection work of ultrathin sections with a single slice thickness of 30 - 50 nm, once the stress between the sample and the diamond knife is released and re-operation is performed, such as knife alignment and block trimming, it will cause slight changes in the knife table. Even for experienced ultrathin section workers, when collecting ultrathin sections again, it will cause instability during the cutting of the starting ultrathin section, manifested as problems such as local damage and uneven thickness of the sections, and 3 - 5 ultrathin sections will be lost. The overall thickness loss is between 90 - 250 nm. This volume represents the information content of 3 - 4 vesicles and synapses in neural circuit reconstruction. In either case, it will cause discontinuity of serial ultrathin sections and loss of intermediate section information.

[0005] At the existing instrument and technology level, the maximum volume of the continuous ultra-thin sections obtained is 200 μm in travel. For life science research, a travel of 200 μm is far from sufficient.

[0006] Accordingly, there is a need in the art for a new ultra-microtome and its knife holder to solve the above problems. Summary of the Invention

[0007] To solve the above problems in the prior art, that is, to solve the problem that the travel of the sections cut by the existing ultra-microtome is short and cannot meet the scientific research needs, the present invention provides a knife holder for an ultra-microtome, and the knife holder includes:

[0008] A knife holder body, and the knife holder is mounted on the base of the ultra-microtome through the knife holder body;

[0009] A tool holder, which is connected to the knife holder body and is used for mounting a target tool;

[0010] A driving device, which is arranged on the knife holder body or between the knife holder body and the tool holder, and the driving device is used for driving the tool holder to move along a preset direction. <(

[0011] In a preferred technical solution of the above knife holder, the knife holder further includes a rotating device arranged between the knife holder body and the tool holder, the rotating device can drive the tool holder to rotate, and the driving device drives the tool holder to move through the rotating device.

[0012] In a preferred technical solution of the above knife holder, the rotating device includes a first rotating member and a second rotating member that are rotatably connected together, the first rotating member is fixedly connected to the tool holder, and the second rotating member is fixedly connected to the knife holder body.

[0013] In a preferred technical solution of the above knife holder, a plurality of arc-shaped tooth-like structures are arranged on one side of the first rotating member facing the second rotating member; a worm that meshes with the tooth-like structures is pivotally arranged on the second rotating member, and when the worm rotates, it can drive the first rotating member to rotate.

[0014] In a preferred technical solution of the above knife holder, at least one end of the worm is provided with an operating handle; and / or, scale lines for marking the rotation angle of the first rotating member are arranged on the first rotating member and / or the second rotating member.

[0015] In a preferred technical solution of the above knife holder, the knife holder further includes a knife holder base that is slidably connected to the knife holder body, and the knife holder is mounted on the base of the ultra-microtome through the knife holder base.

[0016] In the preferred technical solution of the above tool rest, a dovetail groove is provided on the tool rest base, and a convex structure matching the dovetail groove is provided on the tool rest body. The tool rest base and the tool rest body are slidably connected together through the dovetail groove and the convex structure; and / or, a fixing mechanism for fixing the tool rest body to the tool rest base is provided on the tool rest base; and / or, an adjusting mechanism for adjusting the position of the tool rest body on the tool rest base is provided on the tool rest base.

[0017] In the preferred technical solution of the above tool rest, the driving device is a piezoelectric ceramic sheet provided on the tool rest body. When the piezoelectric ceramic sheet is energized, it can generate deformation and thus drive the tool holder to move.

[0018] In the preferred technical solution of the above tool rest, the tool rest body includes a first tool rest body connected to the tool holder and a second tool rest body connected to the base of the ultramicrotome. One end of the piezoelectric ceramic sheet is connected to the first tool rest body, and the other end of the piezoelectric ceramic sheet is connected to the second tool rest body.

[0019] In addition, the present invention also provides an ultramicrotome, characterized in that the ultramicrotome includes the tool rest according to any one of the above preferred embodiments.

[0020] Those skilled in the art can understand that in the preferred technical solution of the present invention, by setting the tool holder to be movable relative to the tool rest body and enabling the tool holder to be driven by a driving device, the tool rest of the present invention can drive the target tool to move along a preset direction, thereby slicing the sample, effectively increasing the slicing stroke of the ultramicrotome and making the stroke of the slices cut by the ultramicrotome longer. Compared with the ultramicrotome that only moves the sample arm in the prior art, the ultramicrotome of the present invention greatly increases the stroke length of slicing and meets the scientific research requirements.

[0021] Further, the driving device is a piezoelectric ceramic sheet provided on the tool rest body.

[0022] Preferably, the piezoelectric ceramic sheet of the present invention is set to generate a deformation of 1 mm when energized, so as to increase the moving distance of the tool holder and make the stroke length of the slice reach 1 mm, thereby meeting the stroke length requirement of slicing.

[0023] Furthermore, the tool rest body includes a first tool rest body and a second tool rest body. The first tool rest body is connected to the tool holder, and the second tool rest body is connected to the base of the ultra-thin press. One end of the piezoelectric ceramic sheet is connected to the first tool rest body, and the other end of the piezoelectric ceramic sheet is connected to the second tool rest body.

[0024] Furthermore, a rotating device is provided between the first tool rest body and the tool holder, and the rotating device is used to adjust the angles of the tool holder and the target tool. The second tool rest body is connected to the base of the ultra-thin tablet press through a tool rest base. Specifically, the second tool rest body is slidably connected to the tool rest base, and the tool rest base is fixedly connected to the base of the ultra-thin tablet press. So that the tool rest body can quickly approach the sample with the tool holder, facilitating the operation of the operator. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] The preferred embodiments of the present invention will be described below with reference to the accompanying drawings, in which:

[0026] Figure 1 is a side view of the tool rest of the ultra-thin microtome of the present invention;

[0027] Figure 2 is a front view of the tool rest of the ultra-thin microtome of the present invention;

[0028] Figure 3 is a top view of the tool rest of the ultra-thin microtome of the present invention;

[0029] Figure 4 is a left view of the tool rest of the ultra-thin microtome of the present invention;

[0030] Figure 5 is a schematic structural diagram of the tool rest body of the present invention;

[0031] Figure 6 is a partial schematic view of the lower tool rest of the tool rest body of the present invention;

[0032] Figure 7 is a schematic connection diagram of the tool rest body of the present invention and the adjustment knob.

[0033] LIST OF REFERENCE NUMERALS:

[0034] 1, diamond knife; 2, tool holder; 3, diamond knife fixing knob; 4, first rotating member; 5, second rotating member; 6, angle adjustment knob; 7, tool rest body; 701, lower tool rest; 7011, chute; 7012, coarse rack; 7013, fine rack; 702, upper tool rest; 8, tool rest fixing knob; 9, tool rest base; 10, first position coarse adjustment knob; 11, second position coarse adjustment knob; 111, coarse adjustment gear; 12, second position fine adjustment knob; 121, fine adjustment gear; 13, piezoelectric ceramic sheet; 14, slider. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0035] Those skilled in the art should understand that the embodiments in this section are only used to explain the technical principles of the present invention and are not used to limit the protection scope of the present invention. For example, although the components in the drawings are drawn according to a certain proportional relationship, this proportional relationship is not fixed. Those skilled in the art can adjust it according to needs to adapt to specific application scenarios, and the adjusted technical solutions will still fall within the protection scope of the present invention.

[0036] It should be noted that in the description of the present invention, the terms indicating directions or positional relationships such as "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer", etc. are based on the directions or positional relationships shown in the drawings. This is only for convenience of description and does not indicate or imply that the device or element must have a specific orientation, be constructed and operated in a specific orientation, so it cannot be understood as a limitation of the present invention. In addition, the terms "first", "second", "third" are only used for descriptive purposes and cannot be understood as indicating or implying relative importance.

[0037] In addition, it should also be noted that in the description of the present invention, unless otherwise clearly specified and limited, the terms "installation", "connection", "connection" should be understood in a broad sense. For example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be directly connected or indirectly connected through an intermediate medium, and it can be the communication inside two elements. For those skilled in the art, the specific meanings of the above terms in the present invention can be understood according to specific situations.

[0038] Although not shown in the figure, the ultramicrotome of the present invention includes a machine body, a sample arm, a base, and a knife table. Among them, the sample arm and the base are respectively fixedly connected to the machine body, and the sample arm is used to place samples. The knife table is fixedly connected to the base. Since the machine body, sample arm, and base of the ultramicrotome are structures and devices well-known to those skilled in the art, no further elaboration will be made here.

[0039] As Figures 1 to 4 shown, the knife table of the present invention mainly includes a knife holder 2, a rotating device (not marked in the figure), a knife table body 7, and a knife table base 9, which are connected in sequence from top to bottom.

[0040] Continue to refer to Figures 1 to 4 , the knife holder 2 is used to install and fix the diamond knife 1 as the target tool. In addition, those skilled in the art can also replace the diamond knife 1 with any other feasible tool according to needs, such as a metal knife, a ceramic knife, a resin knife, etc.

[0041] Although not explicitly shown in the figures, the tool holder 2 of the present invention mainly includes a tool holder body fixedly connected to a rotating device and two clamping pieces slidably disposed on the tool holder body. The two clamping pieces can approach and separate from each other. When the two clamping pieces approach each other, the two clamping pieces can clamp the diamond tool 1 therebetween, and thus fasten the diamond tool 1 to the tool holder 2.

[0042] As Figure 1 shown, the tool rest of the present invention further includes a diamond tool fixing knob 3. The diamond tool fixing knob 3 is used to make the two clamping pieces on the tool holder 2 approach or separate from each other. Specifically, a threaded hole is respectively provided on each of the two clamping pieces of the tool holder 2, and the threads of the two threaded holes have opposite helix directions. The diamond tool fixing knob 3 includes a screw rod and a knob fixedly connected together or integrally formed. Wherein, the external thread on the screw rod matches the internal thread in the two threaded holes. When the diamond tool fixing knob 3 rotates forward / backward, the screw rod drives the two clamping pieces to approach each other; when the diamond tool fixing knob 3 rotates backward / forward, the screw rod drives the two clamping pieces to separate from each other. Preferably, the maximum distance between the two clamping pieces is not less than 12 mm, so that the tool holder 2 can hold the diamond tool 1 with a size of 6 - 12 mm.

[0043] Continuing to refer to Figures 1 to 4 , the rotating device mainly includes a first rotating member 4, a second rotating member 5, and an angle adjustment knob 6. Wherein, the first rotating member 4 is fixedly connected to the tool holder 2, the second rotating member 5 is fixedly connected to the tool rest body 7, and the first rotating member 4 and the second rotating member 5 are pivotally connected together. Exemplarily, a cylinder is provided on one side of the first rotating member 4 close to the second rotating member 5, and a round hole matching the cylinder is provided on the second rotating member 5. The cylinder is inserted into the round hole, so that the first rotating member 4 and the second rotating member 5 are pivotally connected together.

[0044] As Figure 1 and Figure 3 shown, the angle adjustment knob 6 is an example of an adjustment mechanism, which is pivotally disposed on the second rotating member 5, and the angle adjustment knob 6 can drive the first rotating member 4 to rotate when it rotates. Specifically, the angle adjustment knob 6 includes a worm and operating handles located at both ends of the worm. Wherein, the worm and the two operating handles are fixedly connected or integrally formed, and the worm is located inside the second rotating member 5 in Figure 1 , and the two operating handles are located outside the second rotating member 5 in Figure 1 . Those skilled in the art can understand that, on the premise of being able to drive the worm to rotate, only one operating handle can be provided. Further, a plurality of tooth-shaped structures are provided on one side of the first rotating member 4 close to the second rotating member 5 and corresponding to the worm. The plurality of tooth-shaped structures are arranged in an arc shape at equal intervals around the axis of the cylinder, so that the worm can mesh with each tooth-shaped structure.

[0045] In the assembled state of the rotating device, the operator can drive the first rotating member 4, the tool holder 2 and the diamond tool 1 to rotate by rotating the rotation angle adjustment knob 6. In a preferred embodiment of the present invention, the angle range within which the first rotating member 4 can rotate is ±15°. Preferably, when the rotation angle adjustment knob 6 rotates one full turn, the first rotating member 4 rotates 5°. In addition, those skilled in the art can also adjust the angle range within which the first rotating member 4 can rotate to any other arbitrary value as needed, such as ±10°, ±20°, ±30°, etc.; and set the adjustment precision of the rotation angle adjustment knob 6 to any other feasible setting, such as 1° / turn, 1.5° / turn, 2° / turn, 2.5° / turn, etc. Those skilled in the art can understand that the rotation angle adjustment knob 6 can be set to any feasible diameter and length, for example, a diameter of 8.6 mm and a length of 27.7 mm.

[0046] Furthermore, although not shown in the figures, in a preferred embodiment of the present invention, scale lines for marking the rotation angle of the first rotating member 4 are provided on the first rotating member 4 and / or the second rotating member 5. Exemplarily, scale lines are provided on the second rotating member 5 and a pointer is provided on the first rotating member 4.

[0047] As Figure 5 shown, the tool rest body 7 of the present invention includes a lower tool rest 701 and an upper tool rest 702 that are fixedly connected together. Among them, the lower tool rest 701 is slidably connected to the tool rest base 9, and the upper tool rest 702 is fixedly connected to the second rotating member 5. The tool rest of the present invention further includes a piezoelectric ceramic sheet 13 disposed between the lower tool rest 701 and the upper tool rest 702. In a preferred embodiment of the present invention, the piezoelectric ceramic sheet 13 is made of aluminum alloy and has a deformation precision of 5 nm. Or those skilled in the art can also use any other form of piezoelectric ceramic sheet 13 as needed and set it to any other feasible precision. For example, set the deformation precision of the piezoelectric ceramic sheet 13 to 1 nm, 3 nm, 8 nm, etc. Further, the size of the piezoelectric ceramic sheet 13 is 68×40×26.2 mm , Figure 5 , 3 ,

[0048] , Figure 5 . And when the piezoelectric ceramic sheet 13 is energized, its deformation amount in the left - right direction along Figure 5 is at least 1 mm. In addition, those skilled in the art can also set the size of the piezoelectric ceramic sheet 13 to any other arbitrary value as needed, so that the deformation amount of the piezoelectric ceramic sheet 13 in the left - right direction along Figure 5 is also any arbitrary value. For example, make the deformation amount of the piezoelectric ceramic sheet 13 in the left - right direction along Figure 5 at least 500 μm.

[0048] Continue to refer to Figure 5, a first protrusion is provided on one side of the lower tool rest 701 close to the upper tool rest 702, and a second protrusion is provided on one side of the upper tool rest 702 close to the lower tool rest 701. One end of the piezoelectric ceramic sheet 13 abuts against the first protrusion, and the other end of the piezoelectric ceramic sheet 13 also abuts against the second protrusion.

[0049] As Figure 1 and Figure 2 shown, a dovetail groove (not labeled in the figure) is provided on the tool rest base 9, and a protruding structure (not labeled in the figure) matching the dovetail groove is provided on the lower tool rest 701. The lower tool rest 701 is slidably connected to the tool rest base 9 through the protruding structure and the dovetail groove.

[0050] Continue to refer to Figure 1 and Figure 2 , the tool rest of the present invention further includes a tool rest fixing knob 8. As an example of the fixing mechanism, the tool rest fixing knob 8 is used to fix the lower tool rest 701 to any position on the tool rest base 9 to prevent the lower tool rest 701 from sliding. Specifically, the tool rest fixing knob 8 includes a screw rod, and a threaded through hole matching the screw rod is provided on the tool rest base 9. The screw rod is screwed into the threaded through hole. When the tool rest fixing knob 8 is tightened, one end of the tool rest fixing knob 8 abuts against the lower tool rest 701, and thus the lower tool rest 701 is locked to the tool rest base 9.

[0051] As Figures 1 to 3 shown, the tool rest of the present invention further includes an adjusting structure. The adjusting mechanism includes a first position coarse adjustment knob 10, a second position coarse adjustment knob 11, and a second position fine adjustment knob 12. Among them, the first position coarse adjustment knob 10 is used to drive the lower tool rest 701 to move along the X direction ( Figure 3 the left - right direction in Figure 3 ), and the second position coarse adjustment knob 11 and the second position fine adjustment knob 12 are used to drive the lower tool rest 701 to move along the Y direction (

[0052] the up - down direction in Figure 1 ). Figure 6 and 7 shown).

[0053] As Figure 6 and Figure 7 shown, on one side of the lower tool rest 701 ( Figure 3 the right side of the lower tool rest 701 in

[0054] ), a convex - shaped sliding groove 7011 is provided, a slider 14 is arranged in the sliding groove 7011, and a threaded hole is provided on the slider 14. A coarse rack 7012 and a fine rack 7013 are provided on the bottom side of the lower tool rest 701. Figure 7As shown, one end of the first-position coarse adjustment knob 10 extending into the tool rest base 9 is provided with an external thread (not shown in the figure), and this external thread matches the threaded hole on the slider 14. By rotating the first-position coarse adjustment knob 10, the lower tool rest 701 can be moved upward or downward (left or right along the direction shown in Figure 3 ).

[0055] Continue to refer to Figure 7 , the second-position coarse adjustment knob 11 is a hollow rod-shaped structure, which is sleeved on the second-position fine adjustment knob 12, and the second-position coarse adjustment knob 11 and the second-position fine adjustment knob 12 can rotate independently. One end of the second-position coarse adjustment knob 11 extending into the tool rest base 9 is provided with a coarse adjustment gear 111 meshing with the coarse rack 7012, and one end of the second-position fine adjustment knob 12 extending into the tool rest base 9 is provided with a fine adjustment gear 121 meshing with the fine rack 7013. By rotating the second-position coarse adjustment knob 11, the lower tool rest 701 can be quickly moved left or right (upward or downward along the direction shown in Figure 3 ). By rotating the second-position fine adjustment knob 12, the lower tool rest 701 can be slowly and precisely moved left or right (upward or downward along the direction shown in Figure 3 ).

[0056] Next, refer to Figure 1 to describe in detail the working principle of the tool rest of the ultra-thin slicer of the present invention.

[0057] As Figure 1 shown, the steps of using the tool rest of the present invention are as follows:

[0058] First step, first fix the diamond knife 1 to the tool holder 1 through the diamond knife fixing knob 3.

[0059] Second step, manually push the rotating device to make the cutting edge of the diamond knife 1 advance along the direction of the arrow in Figure 1 , and quickly approach the sample arm.

[0060] Third step, by adjusting the second-position coarse adjustment knob 11 and the second-position fine adjustment knob 12, make the cutting edge of the diamond knife 1 close enough to the sample arm, or just touch the sample on the sample arm.

[0061] Fourth step, rotate the first rotating member 4 by the angle adjustment knob 6 to make the cutting edge of the diamond knife 1 parallel to the sample.

[0062] Fifth step, energize the piezoelectric ceramic sheet 13 to generate deformation, drive the upper tool rest 702 to move, and make the diamond knife 1 slice the sample.

[0063] Sixth step, after slicing is completed, cut off the power supply of the piezoelectric ceramic sheet 13 to restore it to its original shape, and make the diamond knife 1 return to its original position.

[0064] Among them, in the fifth step, the piezoelectric ceramic sheet 13 can generate different deformation amounts according to the magnitude of the energizing voltage and / or current. In a preferred embodiment of the present invention, the deformation amount of the piezoelectric ceramic 13 each time is 1 unit length, and the deformation amount of the piezoelectric ceramic 13 is superimposed by increasing the voltage and / or current. Those skilled in the art can understand that the unit length can be any value, such as 5 nm, 15 nm, 20 nm, 30 nm, etc. Those skilled in the art can understand that the diamond knife 1 of the present invention can cut the sample once when the deformation amount of the piezoelectric ceramic 13 increases by 1 unit length each time, or can also cut the sample once when the deformation amount of the piezoelectric ceramic 13 increases by multiple (such as 2, 3, 4, etc.) unit lengths.

[0065] It should be noted that the stroke mentioned in the present invention refers to the deformation amount generated by the piezoelectric ceramic 13. The maximum stroke refers to the maximum deformation amount that the piezoelectric ceramic 13 can generate, that is, the total thickness of the sample section that the diamond knife 1 can cut out.

[0066] In addition, although not shown in the figure, in a preferred embodiment of the present invention, the tool holder 2 is inclined upward by 6° so that the diamond knife 1 can section the sample. Or those skilled in the art can also make the tool holder 2 inclined at any feasible angle according to needs, such as 5°, 8°, 10°, etc. Further, an LED backlight is also provided at the front end ( Figure 3 the upper end of the tool holder body 7 and / or the tool holder base 9) so that the operator can observe the working state of the ultramicrotome. Furthermore, the ultramicrotome of the present invention further includes a controller, and the controller can make an alarm when the diamond knife 1 is about to move to the end of the stroke.

[0067] Based on the above description, those skilled in the art can understand that the tool holder of the ultramicrotome of the present invention can provide a stroke of up to 1 mm for the diamond knife 1. Therefore, the sections of the samples cut by the ultramicrotome of the present invention can reach 1 mm, meeting the requirements of researchers.

[0068] The following are three examples of the use of the ultramicrotome of the present invention.

[0069] Example 1 (>200 μm range ultramicrotome section collection, tool holder driven alone):

[0070] Prepare a sample resin-embedded block, and expose the surface and side of the diamond knife by fine trimming. The size is 0.5×0.8×1 mm (X×Y×Z). After being fixed in the sample clamp, it is stably installed on the sample arm of the microtome.

[0071] Turn on the controller switch, install the knife table, and install a 4mm-edge diamond knife 1 for continuous ultra-thin sectioning. Fix the diamond knife 1 in the knife holder 2 with the diamond knife fixing knob 3, and push the knife table body 7 close enough to the sample surface.

[0072] Turn on the LED backlight on the controller panel and adjust it to an appropriate brightness. Rotate the coarse adjustment knob 11 at the second position and the fine adjustment knob 12 at the second position to push the knife table close enough to the sample surface. Fix the knife table body 7 to the knife table base 9 with the knife table fixing knob 8. Adjust the angle adjustment knob 6 to make the sample surface parallel to the edge of the diamond knife 1, and adjust the sample arm clearance angle and tilt angle so that both the upper and lower edges of the sample are parallel to the edge of the knife, complete the knife alignment, fill with water, install and start the collector.

[0073] Set the sectioning window in the microtome control panel, and set the number of window grids to 3; set the section thickness to 0 so that the microtome only completes the up-and-down swing of the sample arm during operation; set the sectioning speed to 0.5mm / s.

[0074] Set the section thickness to 50nm in the control panel, record the number of sectioning windows as 3, the sectioning speed as 0.5mm / s, and set the knife table stepping direction forward.

[0075] The knife table stepping interval time = (3 * 0.5) / 0.5 + (30 - 3) * 0.5 / 10 = 4.35s. The knife table automatically calculates the knife table stepping interval as 4.35 seconds according to the sectioning window size and sectioning speed.

[0076] Turn the microtome handwheel to the lowest end of the stroke, click the sectioning button on the microtome control panel to start the swing of the sample arm. At the same time, click the sectioning button on the knife table control panel to start the stepping of the knife table, start sectioning, and continuously collect the cut sections.

[0077] When the number of sections in the microtome panel is 6000 and the knife table stepping number displayed on the knife table control panel is 300μm, stop the microtome, stop the knife table, stop the film collector, collect the last section that has not been collected onto the collector, and collect the ultra-thin sections in the water bath onto a silicon wafer with appropriate hydrophilicity with a camel hair brush for atomic force microscope testing of the actual section thickness and surface observation with a field emission scanning electron microscope.

[0078] The atomic force microscope test shows that the actual thickness of the 50nm-thick ultra-thin section driven by the sectioning knife table is 51nm, with a deviation of 2% from the actual set value. No tremor marks that may be caused by knife table stepping were found in the electron microscope observation of the sections.

[0079] Example 2 (collection of 30nm ultra-thin sections in a small range, independent drive of the knife table):

[0080] Prepare a resin-embedded sample block, and use a diamond knife to precisely trim the surface and expose the side. The size is 0.5×0.8×0.2 mm. After fixing it in the sample holder, stably install it on the sample arm of the microtome.

[0081] Turn on the controller switch, install the knife stage, and install a 3-mm-edge diamond knife 1 for continuous ultra-thin sectioning. Fix diamond knife 1 in knife holder 2 with diamond knife fixing knob 3, and push knife stage body 7 close enough to the sample surface.

[0082] Turn on the LED backlight on the controller panel and adjust it to an appropriate brightness. Rotate the coarse adjustment knob 11 in the second position and the fine adjustment knob 12 in the second position to push the knife stage close enough to the sample surface. Fix knife stage body 7 to knife stage base 9 with knife stage fixing knob 8. Adjust angle adjustment knob 6 to make the sample surface parallel to the edge of diamond knife 1. Adjust the sample arm clearance angle and tilt angle so that both the upper and lower sides of the sample are parallel to the edge of the knife to complete knife alignment. Inject water, install and start the collector.

[0083] Set a sectioning window in the microtome control panel. The number of window grids set is 3; set the section thickness to 0 so that the microtome only completes the up-and-down swing of the sample arm during operation; set the sectioning speed to 0.4 mm / s.

[0084] Set the section thickness to 30 nm in the control panel, record the number of sections in the sectioning window as 3, the sectioning speed as 0.4 mm / s, and set the knife stage stepping direction forward.

[0085] Knife stage stepping interval time = (3 * 0.5) / 0.4+(30 - 3)*0.5 / 10 = 5.1 s. The knife stage automatically calculates the knife stage stepping interval as 5.1 seconds according to the sectioning window size and sectioning speed.

[0086] Turn the microtome handwheel to the lowest end of the stroke, click the sectioning button on the microtome control panel to start the swing of the sample arm. At the same time, click the sectioning button on the knife stage control panel to start the stepping of the knife stage, start sectioning, and continuously collect the cut sections.

[0087] When the number of sections in the microtome panel is 500 and the knife stage stepping number displayed on the knife stage control panel is 15 μm, stop the microtome, stop the knife stage, stop the section collector, collect the last section that has not been collected onto the collector, and use a camel hair brush to collect the ultra-thin sections in the water bath onto a silicon wafer with appropriate hydrophilicity for atomic force microscope testing of the actual section thickness and field emission scanning electron microscope for surface observation.

[0088] The actual thickness of the 30-nm-thick ultra-thin section driven by the sectioning knife stage measured by the atomic force microscope is 30.6 nm, with a deviation of 2% from the actual set value. No tremor marks that may be caused by knife stage stepping were found in the electron microscope observation of the sections.

[0089] Example 3: (1mm ultra-long range ultra-thin section collection, simultaneous drive of sample arm and knife table)

[0090] Prepare the sample resin-embedded block, finely trim the surface and expose the side with a diamond knife, with a size of 0.5×1×1.2mm. After fixing it in the sample clamp, stably install it on the sample arm of the microtome.

[0091] Turn on the controller switch, install the knife table, and install a 3mm-edge diamond knife for continuous ultra-thin sectioning. Fix the diamond knife 1 in the knife holder 2 with the diamond knife fixing knob 3, and push the knife table body 7 close enough to the sample surface.

[0092] Turn on the LED backlight on the controller panel and adjust it to the appropriate brightness. Rotate the coarse adjustment knob 11 in the second position and the fine adjustment knob 12 in the second position to push the knife table close enough to the sample surface. Fix the knife table body 7 to the knife table base 9 through the knife table fixing knob 8. Adjust the angle adjustment knob 6 to make the sample surface parallel to the edge of the diamond knife 1. Adjust the sample arm clearance angle and tilt angle so that both the upper and lower edges of the sample are parallel to the edge of the knife, complete the knife alignment, inject water, install and start the collector.

[0093] Set the sectioning window in the microtome control panel, with the number of window grids set to 4; set the section thickness to 20, so that the microtome completes the up and down swing of the sample arm and the sectioning step when running; set the sectioning speed to 1mm / s.

[0094] Set the section thickness to 80nm in the control panel, record the number of sectioning windows as 4, the sectioning speed as 1mm / s, and set the knife table stepping direction forward.

[0095] The knife table stepping interval time = (4*0.5) / 1+(30 - 4)*0.5 / 10 = 3.3s. The knife table automatically calculates the knife table stepping interval as 3.3 seconds according to the sectioning window size and sectioning speed.

[0096] Turn the microtome handwheel to the lowest end of the stroke, click the sectioning button on the microtome control panel, and start the swing of the sample arm. At the same time, click the sectioning button on the knife table control panel, start the stepping of the knife table, start sectioning, and continuously collect the cut sections.

[0097] When the number of sections in the microtome panel is 10000 and the number of knife table steps displayed in the knife table control panel is 800μm, at this time, the number of sections recorded in the microtome is 10000, the number of steps of the microtome sample arm is 200μm. Stop the microtome, stop the knife table, stop the section collector. Use a camel hair brush to collect the ultra-thin sections that were not finally collected into the collector and were in the water tank onto a silicon wafer with appropriate hydrophilicity, and perform atomic force microscope testing on the actual thickness of the sections and surface observation with a field emission scanning electron microscope.

[0098] The atomic force microscope test was driven by the sample arm of the microtome and the knife table drive. The actual thickness of the ultra-thin section with a thickness of 100 nm was 101 nm, and the deviation from the actual set value was 1%. No tremor marks that might be caused by the knife table stepping were found by electron microscopy observation of the section.

[0099] So far, the technical solution of the present invention has been described in conjunction with the preferred embodiments shown in the accompanying drawings. However, it is easy for those skilled in the art to understand that the protection scope of the present invention is obviously not limited to these specific embodiments. Without departing from the principle of the present invention, those skilled in the art can make equivalent changes or substitutions to the relevant technical features, and the technical solutions after these changes or substitutions will fall within the protection scope of the present invention.

Claims

1. A knife rest of an ultramicrotome, characterized in that, The tool carrier includes: A tool carrier body, and the tool carrier is mounted on the base of the ultramicrotome through the tool carrier body; A tool rest, which is connected to the tool carrier body and is used for mounting a target tool; A driving device, which is arranged on the tool carrier body or between the tool carrier body and the tool rest, and the driving device is used for driving the tool rest to move along a preset direction; Two clamping pieces, the two clamping pieces are slidably arranged on the tool rest body of the tool rest, the two clamping pieces can approach and move away from each other, and when the two clamping pieces approach, the two clamping pieces can clamp the target tool therebetween; Wherein, the driving device is a piezoelectric ceramic sheet arranged on the tool carrier body, and when the piezoelectric ceramic sheet is energized, it can generate different deformation amounts according to the magnitude of the energization voltage and / or current, and thus drive the tool rest to move; and The maximum deformation amount generated by the piezoelectric ceramic sheet is the total thickness of the sample section that the target tool can cut out.

2. The knife rest of the ultra-thin slicer according to claim 1, characterized in that, The tool carrier further includes a rotating device arranged between the tool carrier body and the tool rest, and the rotating device can drive the tool rest to rotate, The driving device drives the tool rest to move through the rotating device.

3. The knife rest of the ultra-thin slicer according to claim 2, characterized in that, The rotating device includes a first rotating member and a second rotating member that are rotatably connected together, the first rotating member is fixedly connected to the tool rest, and the second rotating member is fixedly connected to the tool carrier body.

4. The knife rest of the ultra-thin slicer according to claim 3, characterized in that, A plurality of tooth-shaped structures arranged in an arc are provided on one side of the first rotating member facing the second rotating member; A worm meshing with the tooth-shaped structure is pivotally arranged on the second rotating member, and when the worm rotates, it can drive the first rotating member to rotate.

5. The knife rest of the ultra-thin slicer according to claim 4, characterized in that, At least one end of the worm is provided with an operating handle; And / or, scale lines for marking the rotation angle of the first rotating member are provided on the first rotating member and / or the second rotating member.

6. The knife rest of the ultra-thin slicer according to claim 1, characterized in that, The tool carrier further includes a tool carrier base slidably connected to the tool carrier body, and the tool carrier is mounted on the base of the ultramicrotome through the tool carrier base.

7. The knife rest of the ultra-thin slicer according to claim 6, characterized in that, A dovetail groove is provided on the tool carrier base, and a convex structure matching the dovetail groove is provided on the tool carrier body, and the tool carrier base and the tool carrier body are slidably connected together through the dovetail groove and the convex structure; And / or, a fixing mechanism for fixing the tool carrier body to the tool carrier base is provided on the tool carrier base; And / or, an adjusting mechanism for adjusting the position of the tool carrier body on the tool carrier base is provided on the tool carrier base.

8. The knife table of the ultra-thin slicer according to claim 1, characterized in that, The tool carrier body includes a first tool carrier body connected to the tool rest and a second tool carrier body connected to the base of the ultramicrotome, one end of the piezoelectric ceramic sheet is connected to the first tool carrier body, and the other end of the piezoelectric ceramic sheet is connected to the second tool carrier body.

9. An ultramicrotome, characterized in that, The ultramicrotome includes the tool carrier according to any one of claims 1 to 8.

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