Full-automatic high-flux electron microscope grid dyeing machine

Abstract

The invention provides a full-automatic high-flux electron microscope carrying net dyeing machine which comprises a conveying belt used for bearing a copper net sample, a visual collecting device arranged above the conveying belt and a coloring agent spraying device, the coloring agent spraying device is provided with a liquid spraying trolley capable of moving in the X direction and the Y direction, the liquid spraying trolley is located above the conveying belt, and the liquid spraying trolley is located above the conveying belt. The liquid spraying trolley is located at the downstream of the visual collection device, is connected with the coloring agent box through a flexible pipe, and is used for spraying a coloring agent on the copper wire mesh sample according to the position of the copper wire mesh sample collected by the visual collection device and the running speed of the conveying belt. According to the spraying device, accurate coloring agent spraying can be achieved, and after the first coloring agent is sprayed, only the copper net needs to be cleaned. Through image acquisition and automatic tracking of the liquid spraying trolley, placement of copper mesh samples is not limited by position precision, the cross arm sampling device arranged at the tail of the conveying belt can automatically take the dyed samples into the lifting type tray, the labor intensity is greatly reduced, and the efficiency of large-scale and large-quantity dyeing operation is greatly improved.

Description

Technical Field

[0001] The invention relates to the technical field of biological sample staining, in particular to a full-automatic high-throughput electron microscope grid staining machine. Background Art

[0002] The observation and image analysis of samples by transmission electron microscopy (TEM) include the following steps: 1. Sample collection: collect appropriate tissue samples; 2. Fixation: fix with chemical reagents; 3. Dehydration: remove moisture from the sample; 4. Infiltration and embedding: put the dehydrated sample into the resin for infiltration, and then solidify to form a solid block; 5. Ultrathin sectioning: use an ultramicrotome to cut the embedded block to obtain slices with a thickness of about 50-100 nanometers. 6. Staining: stain the slices loaded on the copper mesh. 7. Microscopic observation. The current staining method uses manual operation, which is to make the staining liquid into water droplets, and then flip the copper mesh close to the water droplets to stain the slices on the copper mesh. The problem is that some staining liquids need to be immersed for a long time, such as 10 minutes. During this process, the slices may fall off, resulting in the failure of copper mesh sample preparation, and the technical requirements for operators are high. Especially in the case of large-scale sample experiments, the existing operation method is too inefficient, affecting the efficiency of the experiment. Chinese patent document CN217820089U records an integrated device for clamping and dyeing copper mesh for electron microscope, which uses a lead screw as a driving mechanism and clamps the copper mesh with tweezers. This solution does not improve the efficiency of dyeing. CN210376190U records an electron microscope staining sample preparation device, which uses a solution of top dripping to dye the copper mesh sample, but this solution has the problem of high difficulty in accurately controlling the amount of dripping. In addition, the document also records the problem that the copper mesh is small and easily rolls over under the influence of dripping. The solution of this document is to set a fixed protrusion 35 to prevent the copper mesh from rolling over, but this structure increases the difficulty of automated operation. Summary of the invention

[0003] The technical problem to be solved by the present invention is to provide a fully automatic high-throughput electron microscope grid staining machine, which can conveniently set the staining agent for the copper grid sample and realize the automation of the staining operation to cope with the substantial increase in experimental samples.

[0004] In order to solve the above-mentioned technical problems, the technical solution of the present invention is: a fully automatic high-throughput electron microscope grid staining machine, comprising a conveyor belt for carrying copper mesh samples, a visual acquisition device is provided above the conveyor belt, and a dye spraying device is also provided. The dye spraying device is provided with a liquid spraying trolley that can move along the X and Y directions. The liquid spraying trolley is located above the conveyor belt, and the liquid spraying trolley is located downstream of the visual acquisition device. The liquid spraying trolley is connected to the dye box through a flexible tube, and the liquid spraying trolley is used to spray the dye on the copper mesh sample according to the position of the copper mesh sample collected by the visual acquisition device and the running speed of the conveyor belt.

[0005] In a preferred embodiment, the copper mesh sample is loaded on a bracket, and the structure of the bracket is as follows: upper pressing plates are provided on both sides of the upper part of the frame, the upper pressing plates are used to clamp the copper mesh sample, the upper pressing plates are pressed on both sides of the copper mesh sample, and a hollow structure is provided at the position corresponding to the frame and the copper mesh sample.

[0006] In a preferred solution, an extended position is further provided on one side of the frame, and a clamping area and an information area are provided on the edge of the extended position.

[0007] In a preferred solution, a second dye spraying device is further provided downstream of the first dye spraying device; An atomizing spray device is provided between the first dye spray device and the second dye spray device; The atomizing spray device cleans the surface of the copper mesh from one side of the copper mesh sample.

[0008] In the preferred solution, the dye spraying device is provided with a transverse guide rail, a transverse beam moving along the transverse guide rail is provided on the transverse guide rail, a liquid spraying trolley moving along the transverse beam is provided on the transverse beam, and the moving range of the liquid spraying trolley covers the entire width of the conveyor belt.

[0009] In the preferred solution, the visual acquisition device acquires the position (x0, y0) of the copper mesh sample; obtains the current coordinates (x j ,y j ), tracking the shortest time (t n ) and the intersection position (x n ,y n ); Output command: The spraying car moves (x n -x j ) absolute value; the transverse beam moves (y n -y j ) absolute value, the spraying car passes through time (t n ) and then start spraying the dye.

[0010] In a preferred solution, if the liquid spraying vehicle needs to continue spraying for multiple times, an instruction is outputted, and the liquid spraying vehicle moves at the same speed as the conveyor belt and continues spraying for a period of time.

[0011] In the preferred solution, if the liquid spraying vehicle needs to continue spraying for multiple times and cover a preset range, an instruction is output and the liquid spraying vehicle generates a motion trajectory of the liquid spraying process so that the liquid spraying vehicle follows the copper mesh sample and reciprocates to cover the copper mesh sample and spray liquid.

[0012] In a preferred solution, a cover is provided at the middle section of the conveyor belt, the cover covers the first dye spraying device and the second dye spraying device, and openings for the bracket to pass through are provided at both ends of the cover; The cover body is a light-shielding structure. The first dye spraying device is separated from the second dye spraying device. Nitrogen is introduced into the cover body space of the second dye spraying device to keep the nitrogen at a slightly positive pressure to prevent oxygen from entering.

[0013] In the preferred solution, a lifting pallet is also provided on one side of the tail end of the conveyor belt. The structure of the lifting pallet is as follows: a pallet rack is slidably installed on a guide rail bracket, one end of the lifting device is connected to the guide rail bracket, and the other end of the lifting device is connected to the pallet rack, so that one layer of the pallet rack is flush with the conveyor belt and the bracket is loaded.

[0014] In a preferred solution, a cross arm sampling device is further provided on one side of the guide rail bracket, and the structure of the cross arm sampling device is as follows: the guide rod is swingably connected to the guide rail bracket via a rotating shaft; The rotating shaft is provided with damping to delay the swing of the guide rail bracket; The guide rod is slidably connected with the sliding rod, and a material taking rod is provided at one end of the sliding rod facing the conveyor belt, and a driving device is also provided to drive the sliding rod to extend and retract; A water absorbing strip is provided on the material taking rod at the same height as the side of the copper mesh sample; A counterweight is provided at one end of the guide rod away from the conveyor belt; When the slide bar is retracted, the pick-up bar is lifted, and when the slide bar is extended, the pick-up bar falls on the surface of the conveyor belt.

[0015] The present invention provides a fully automatic high-throughput electron microscope grid staining machine, which has the following beneficial effects: 1. The present invention can realize automatic and accurate dye spraying on a copper mesh with a thickness of only 1 to 2 mm. After the first dye is sprayed, only the copper mesh needs to be cleaned.

[0016] 2. The present invention uses image acquisition and automatic tracking of the liquid spraying trolley to ensure that the placement of the copper mesh sample is not limited by position accuracy. The horizontal arm sampling device arranged at the tail of the conveyor belt can automatically take the dyed samples into the lifting tray, which greatly reduces labor intensity and greatly improves the efficiency of large-scale and large-quantity dyeing operations.

[0017] 3. The bracket provided in the present invention can prevent the copper mesh sample from turning over during the dyeing process and from falling due to its small size. In particular, the bracket is provided with an information area that can carry the information of the sample, thereby ensuring that the information of the sample is linked to the real object. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The present invention will be further described below in conjunction with the accompanying drawings and embodiments: Figure 1 It is a top view of the overall structure of the present invention.

[0019] Figure 2 It is a schematic diagram of the action of the cross arm sampling device of the present invention.

[0020] Figure 3 It is a schematic structural diagram of the lifting tray of the present invention.

[0021] Figure 4 It is a schematic structural diagram of the bracket of the present invention when loading a copper mesh sample.

[0022] In the figure: bracket 1, clamping area 101, information area 102, upper pressing plate 103, frame 104, conveyor belt 2, cover body 3, dye box 4, flexible tube 5, liquid spray trolley 6, transverse beam 7, transverse guide rail 8, cross arm sampling device 9, material picking rod 91, sliding rod 92, counterweight 93, guide rod 94, material picking motor 95, material picking belt 96, rotating shaft 97, water absorption strip 98, copper mesh sample 10, support platform 11, lifting tray 12, lifting device 121, tray rack 122, guide rail bracket 123, visual collection device 13, second dye spraying device 14, atomizing spray device 15. DETAILED DESCRIPTION

[0023] like Figure 1 In the invention, a fully automatic high-throughput electron microscope grid staining machine includes a conveyor belt 2 for carrying a copper mesh sample 10, a visual acquisition device 13 is provided above the conveyor belt 2, and a dye spraying device is also provided. The dye spraying device is provided with a liquid spraying trolley 6 which can move along the X and Y directions, the liquid spraying trolley 6 is located above the conveyor belt 2, the liquid spraying trolley 6 is located downstream of the visual acquisition device 13, the liquid spraying trolley 6 is connected to the dye box 4 through a flexible tube 5, and the liquid spraying trolley 6 is used to spray the dye on the copper mesh sample 10 according to the position of the copper mesh sample 10 collected by the visual acquisition device 13 and the running speed of the conveyor belt 2.

[0024] The preferred solution is Figure 4 In the figure, the copper mesh sample 10 is loaded on a bracket 1, and the structure of the bracket is as follows: upper pressing plates 103 are provided on both sides above a frame 104, the upper pressing plates 103 are used to clamp the copper mesh sample 10, the upper pressing plates 103 are pressed on both sides of the copper mesh sample 10, and a hollow structure is provided at the position corresponding to the frame 104 and the copper mesh sample 10.

[0025] The preferred solution is Figure 4 In the embodiment, an extended position is provided on one side of the frame 104, and a clamping area 101 and an information area 102 are provided on the edge of the extended position.

[0026] The preferred solution is Figure 1 In the method, a second dye spraying device is further provided downstream of the first dye spraying device; An atomizing spray device 14 is provided between the first dye spray device and the second dye spray device; The first dye spraying device uses uranyl acetate dye, which is used to bind to nucleic acids in cells and increase the electron density of nucleic acids, so that the image under the electron microscope is clearer, which is convenient for observing the distribution and structure of nucleic acids in cells. It is also used to bind to most molecules in cells to increase the contrast of protein and connective tissue fiber components. The second dye spraying device uses lead citrate dye, which is a lead-containing heavy metal salt. Lead ions can specifically bind to certain components in biological samples. Based on the strong scattering ability of heavy metals for electrons, it can enhance the contrast of images and improve the clarity of observation.

[0027] The atomizing spray device 14 cleans the surface of the copper mesh sample 10 from one side thereof. This is to clean the dye and prevent the sample slices from falling off. Specifically, when the copper mesh sample 10 is about 10 to 5 cm away from the atomizing spray device 14, the atomizing spray device 14 starts spraying to clean the dye on the copper mesh sample 10. The atomizing spray device 14 is precisely controlled to prevent a large amount of droplets from falling on the conveyor belt 2.

[0028] The preferred solution is Figure 1 In the figure, the dye spraying device is provided with a transverse guide rail 8, on which is a transverse beam 7 that moves along the transverse guide rail 8, and on the transverse beam 7 is provided a liquid spraying trolley 6 that moves along the transverse beam 7, and the moving range of the liquid spraying trolley 6 covers the entire width of the conveyor belt 2.

[0029] The preferred solution is Figure 1 In the process, the visual acquisition device 13 acquires the position (x0, y0) of the copper mesh sample 10; obtains the current coordinate (x j ,y j), tracking the shortest time (t n ) and the intersection position (x n ,y n ); Output command: Spraying car 6 moves (x n -x j ) absolute value; the transverse beam 7 moves (y n -y j ) absolute value, the spraying car 6 after a certain time (t n ) and then start spraying the dye.

[0030] In a preferred solution, if the liquid spraying trolley 6 needs to continue spraying for multiple times, an instruction is outputted, and the liquid spraying trolley 6 moves at the same speed as the conveyor belt 2 and continues to spray for a period of time.

[0031] In the preferred solution, if the liquid spraying trolley 6 needs to continue spraying for multiple times and cover a preset range, an instruction is output, the liquid spraying trolley 6 generates a motion trajectory of the liquid spraying process, and superimposes the motion trajectory with the motion direction of the conveyor belt 2, so that the liquid spraying trolley 6 follows the copper mesh sample 10 and reciprocates to cover the copper mesh sample 10. The liquid spraying process of the liquid spraying trolley 6 involves "S"-shaped walking, so that the dye is evenly sprayed on the surface of the copper mesh sample 10, and the motion trajectory of the liquid spraying trolley 6 needs to be superimposed with the motion trajectory of the conveyor belt 2.

[0032] The preferred solution is Figure 1 In the embodiment, a cover 3 is provided at the middle of the conveyor belt 2, and the cover 3 covers the entire first and second dye spraying devices, and openings for the bracket 1 to pass through are provided at both ends of the cover 3. The cover 3 is a light-shielding structure, and the first dye spraying device is separated from the second dye spraying device. Nitrogen is introduced into the space of the cover 3 of the second dye spraying device to keep the nitrogen at a slightly positive pressure to prevent oxygen from entering.

[0033] The preferred solution is Figure 1 , 3 In the figure, a lifting pallet 12 is also provided on one side of the tail end of the conveyor belt 2. The structure of the lifting pallet 12 is as follows: a pallet rack 122 is slidably mounted on a guide rail bracket 123, one end of a lifting device 121 is connected to the guide rail bracket 123, and the other end of the lifting device 121 is connected to the pallet rack 122, so that one layer of the pallet rack 122 is flush with the conveyor belt 2, and the bracket 1 is loaded.

[0034] The preferred solution is Figure 1 , 2 In the embodiment, a cross arm sampling device 9 is also provided on one side of the guide rail bracket 123. The structure of the cross arm sampling device 9 is as follows: a guide rod 94 is swingably connected to the guide rail bracket 123 via a rotating shaft 97; The rotating shaft is provided with damping so that the swing of the guide rail bracket 123 has a delay; The guide rod 94 is slidably connected to the sliding rod 92. A material taking rod 91 is provided at one end of the sliding rod 92 facing the conveyor belt 2, and a driving device is also provided to drive the sliding rod 92 to extend and retract. A counterweight 93 is provided at one end of the guide rod 94 away from the conveyor belt 2; A water absorbing strip 98 is provided on the material taking rod 91 at the same height as the side of the copper mesh sample 10. With this structure, when the material taking rod 91 is close to the copper mesh sample 10, the water absorbing strip 98 absorbs excess liquid on the copper mesh sample 10 from the side. After a period of use, the water absorbing strip 98 is replaced.

[0035] When the sliding rod 92 is retracted, the material picking rod 91 is lifted, and when the sliding rod 92 is extended, the material picking rod 91 falls on the surface of the conveyor belt 2.

[0036] like Figure 1In the present invention, when in use, the copper mesh loaded with ultra-thin slices is loaded on the bracket 1, and the bracket 1 is transferred to the conveyor belt 2 in turn by a tray. When the bracket 1 passes under the visual acquisition device 13, the position below the visual acquisition device 13 is set as the coordinate zero point. The visual acquisition device 13 identifies the copper mesh sample 10 on the bracket 1, and sets the center point of the copper mesh sample 10 as the current bracket 1 coordinate, and sets a speed value for the bracket 1. The first dye spraying device calculates the motion trajectory of the liquid spraying trolley 6 according to the coordinate and speed value of the bracket 1. The liquid spraying trolley 6 follows the copper mesh sample 10 according to the motion trajectory and sprays the dye on the surface of the copper mesh sample 10. When passing through the spray atomization device 15, the spray atomization device 15 sprays droplets from the upper side to spray the copper mesh sample 10. The excess dye on the surface of the web sample 10 is removed, and then the second dye is sprayed at the position of the second dye spraying device 14. After the spraying is completed, the multiple brackets 1 are transported to the tail of the conveyor belt 2. At this time, the lifting device 121, preferably a cylinder or an electric push rod, moves to make the tray rack 122 rise or fall an empty tray to align with the height of the conveyor belt 2, and the material taking motor 95 is driven to drive the material taking belt 96 to drive the driven wheel to rotate. The driven wheel is provided with teeth, and the bottom of the sliding rod 92 is provided with teeth engaged with the driven wheel, so as to drive the sliding rod 92 to extend from the guide rod 94. In the initial state, the sliding rod 92 is in a retracted state. Under the action of the counterweight 93, the entire sliding rod 92 is in an inclined state, and the material taking rod 91 leaves the surface of the conveyor belt 2. As the sliding rod 92 extends, the material taking rod 91 falls to a position close to the edge of the conveyor belt 2. Then the material taking motor 95 moves in the reverse direction, and the sliding rod 92 is retracted through the material taking belt 96 and the driven wheel, and the dyed multiple brackets 1 are taken to the empty tray. In the process of taking the brackets 1 back, the water absorption strip 98 absorbs the excess dye on the surface of the copper mesh sample 10, and the lifting tray 12 is raised or lowered by one layer, so that the empty tray is on standby, and the process of fully automatic dyeing of the copper mesh sample 10 is completed. As an important part of the fully automatic production process of ultra-thin sample slices, the present invention lays a foundation for the full automation of the entire ultra-thin sample preparation.

[0037] The above-mentioned embodiments are only preferred technical solutions of the present invention and should not be regarded as limitations of the present invention. The embodiments and features in the embodiments of the present application can be arbitrarily combined with each other without conflict. The protection scope of the present invention shall be the technical solutions recorded in the claims, including the equivalent replacement solutions of the technical features in the technical solutions recorded in the claims. That is, equivalent replacement improvements within this scope are also within the protection scope of the present invention.

Claims

1. A fully automatic high-throughput electron microscope grid staining machine, characterized by: The invention comprises a conveyor belt (2) for carrying a copper mesh sample (10), a visual collection device (13) being arranged above the conveyor belt (2), and a first dye spraying device being arranged, the dye spraying device being provided with a liquid spraying trolley (6) movable in the X and Y directions, the liquid spraying trolley (6) being located above the conveyor belt (2), the liquid spraying trolley (6) being located downstream of the visual collection device (13), the liquid spraying trolley (6) being connected to a dye box (4) via a flexible tube (5), and the liquid spraying trolley (6) being used to spray the dye on the copper mesh sample (10) according to the position of the copper mesh sample (10) collected by the visual collection device (13) and the running speed of the conveyor belt (2).

2. The fully automatic high-throughput electron microscope grid staining machine according to claim 1, characterized in that: The copper mesh sample (10) is loaded on a bracket (1), and the bracket has the following structure: upper pressing plates (103) are provided on both sides above the frame (104), the upper pressing plates (103) are used to clamp the copper mesh sample (10), the upper pressing plates (103) are pressed on the positions on both sides of the copper mesh sample (10), and a hollow structure is provided at the position corresponding to the frame (104) and the copper mesh sample (10); An extended position is also provided on one side of the frame (104), and a clamping area (101) and an information area (102) are provided at the edge of the extended position.

3. The fully automatic high-throughput electron microscope grid staining machine according to claim 2 is characterized in that: A second dye spraying device is also provided downstream of the first dye spraying device; An atomizing spray device (14) is provided between the first dye spray device and the second dye spray device; The atomizing spray device (14) cleans the surface of the copper mesh from one side of the copper mesh sample (10).

4. The fully automatic high-throughput electron microscope grid staining machine according to claim 2 or 3, characterized in that: The dye spraying device is provided with a transverse guide rail (8), a transverse beam (7) moving along the transverse guide rail (8) is provided on the transverse guide rail (8), and a liquid spraying trolley (6) moving along the transverse beam (7) is provided on the transverse beam (7), and the moving range of the liquid spraying trolley (6) covers the entire width of the conveyor belt (2).

5. The fully automatic high-throughput electron microscope grid staining machine according to claim 4, characterized in that: The visual acquisition device (13) acquires the position (x0, y0) of the copper mesh sample (10); obtains the current coordinate (x j ,y j ), tracking the shortest time (t) from the liquid spraying trolley (6) to the intersection of the current copper mesh sample (10). n ) and the intersection position (x n ,y n ); Output command: Spraying trolley (6) moves (x n -x j ) absolute value; the transverse beam (7) moves (y n -y j ), the spraying car (6) after a period of time (t n ) and then start spraying the dye.

6. The fully automatic high-throughput electron microscope grid staining machine according to claim 5, characterized in that: If the liquid spraying trolley (6) needs to continue spraying for multiple times, an instruction is outputted, and the liquid spraying trolley (6) moves at the same speed as the conveyor belt (2) and continues spraying for a period of time.

7. The fully automatic high-throughput electron microscope grid staining machine according to claim 5, characterized in that: If the liquid spraying trolley (6) needs to continue spraying for multiple times and cover a preset range, an instruction is outputted, and the liquid spraying trolley (6) generates a motion trajectory of the liquid spraying process, so that the liquid spraying trolley (6) follows the copper mesh sample (10) and reciprocates to cover the copper mesh sample (10) to spray liquid.

8. The fully automatic high-throughput electron microscope grid staining machine according to claim 1, characterized in that: A cover body (3) is provided at the middle section of the conveyor belt (2), the cover body (3) covers the first dye spraying device and the second dye spraying device, and openings for the bracket (1) to pass through are provided at both ends of the cover body (3); The cover (3) is a light-shielding structure, the first dye spraying device is separated from the second dye spraying device, and nitrogen is introduced into the space of the cover (3) of the second dye spraying device to maintain the nitrogen at a slightly positive pressure to prevent oxygen from entering.

9. The fully automatic high-throughput electron microscope grid staining machine according to any one of claims 1 to 8, characterized in that: A lifting pallet (12) is also provided on one side of the rear end of the conveyor belt (2). The lifting pallet (12) has the following structure: a pallet frame (122) is slidably mounted on a guide rail bracket (123), one end of a lifting device (121) is connected to the guide rail bracket (123), and the other end of the lifting device (121) is connected to the pallet frame (122), so that one layer of the pallet frame (122) is flush with the conveyor belt (2) and the bracket (1) is loaded.

10. The fully automatic high-throughput electron microscope grid staining machine according to claim 9, characterized in that: A cross arm sampling device (9) is also provided on one side of the guide rail bracket (123). The cross arm sampling device (9) has the following structure: a guide rod (94) is swingably connected to the guide rail bracket (123) via a rotating shaft (97); The rotating shaft is provided with damping so that the swinging of the guide rail bracket (123) has a delay; The guide rod (94) is slidably connected to the sliding rod (92); a material taking rod (91) is provided at one end of the sliding rod (92) facing the conveyor belt (2); and a driving device is also provided to drive the sliding rod (92) to extend and retract; A water absorbing strip (98) is provided on the material taking rod (91) at a position equal to the height of the side of the copper mesh sample (10); A counterweight (93) is provided at one end of the guide rod (94) away from the conveyor belt (2); When the sliding rod (92) is retracted, the material picking rod (91) is lifted, and when the sliding rod (92) is extended, the material picking rod (91) falls on the surface of the conveyor belt (2).

Images