Target micro-area laser cutting and collecting device and method for tissue slice on blade

Abstract

The invention discloses a target micro-area laser cutting and collecting device and method for tissue slices on a blade, the target micro-area laser cutting and collecting device for the tissue slices on the blade comprises a carrier table, the blade, a laser cutting system, a control module and a collector, the laser cutting system comprises a laser, a beam expander, a galvanometer and an objective lens, and the scanning angle theta of the galvanometer is adjusted, and the scanned laser is focused in combination with the objective lens, so that the target micro-area on the tissue slice is cut and bounced up. According to the invention, laser microdissection and mechanical slicing are combined, and non-contact collection of a specific micro-area of a tissue slice placed on an inclined blade is realized under the action of laser edge ejection combined with dielectrophoretic force, so that the integration of the form of a sample in the whole acquisition process is ensured, and three-dimensional position information of a target cell or a cell population is accurately reserved; the cutting efficiency is improved, non-contact cutting can be achieved, and samples are prevented from being polluted.

Description

Technical Field

[0001] The present invention relates to the field of optoelectronic imaging, and particularly to a device and method for laser cutting and collecting target micro-regions of tissue sections on a blade. Background Art

[0002] The analysis of spatial omics information of heterogeneous cells or cell populations is of great significance in biomedical research. The isolation of specific cells in biological tissues is the basis for biologists to study the gene expression of heterogeneous cells and further understand biological functions or disease mechanisms. Therefore, a method with high precision and no pollution is needed to separate target cells or cell populations from surrounding biological tissues. Laser microdissection is a powerful technique that can image tissue sections, locate target cells, and achieve high-precision separation using a micro-laser beam, effectively combining morphology and various molecular analyses.

[0003] However, before the existing laser microdissection techniques are implemented, tissue blocks need to be cut into tissue sections by a microtome first, and then cut using a laser microdissection system. For example, the invention patent with the application publication number CN114923755A provides a method for separating Chinese cabbage tissue cells based on laser microdissection technology. After preprocessing the sample, tissue sections are made, and then the processed tissue sections are transferred to the stage of a laser microdissection microscope. The target cells are cut using a laser, and then the samples are collected with a centrifuge tube cap. Usually, when the tissue sections are transferred to the stage of the laser microdissection microscope, the tissue sections also need to be processed into a suitable size for pasting, and then laser cutting operations are performed. This makes it difficult to accurately obtain the three-dimensional position information of target cells or cell populations, and the efficiency of obtaining target micro-regions is relatively low, which has a certain impact on downstream analyses such as RNA and proteins of target micro-regions. At the same time, when transferring tissue sections and collecting target cells, the samples are easily contacted and contaminated. Also, in this type of method, the collection of target cells is carried out manually. In the case where multiple target cells need to be classified and collected, the collection efficiency is low and the samples are easily contaminated. Summary of the Invention

[0004] Therefore, to solve the above problems, the present invention provides a device and method for laser cutting and collecting target micro-regions of tissue sections on a blade.

[0005] The present invention is achieved through the following technical solutions: A device for laser cutting and collecting target micro-regions of tissue sections on a blade, comprising: A carrier stage for placing a sample tissue block and controlling its movement; The blade is inclinedly arranged on one side of the carrier stage through a fixture. The inclination angle of the blade is φ (please provide an angle range and explain the reason / beneficial effect for setting this angle). The blade remains stationary, and the mechanical sectioning is completed by moving the carrier stage to make the blade move relative to the sample tissue block on the carrier stage, thereby obtaining tissue sections. After the cutting is completed, the tissue sections are located on the upper surface of the blade, and the blade serves as the base of the tissue sections; A laser cutting system, including a laser for providing pulsed laser, a beam expander for expanding the laser emitted by the laser, a galvanometer for controlling the scanning angle and scanning speed of the expanded laser, and an objective lens for focusing the scanned laser beam. By adjusting the scanning angle θ of the galvanometer and combining with the objective lens, the scanned laser is focused to achieve the cutting of tissue sections and target micro-regions on the tissue sections and the bouncing of the target micro-regions; A control module is respectively connected to the carrier stage and the laser cutting system in signal, and synchronously controls mechanical sectioning and laser cutting; A collector is located above the target micro-region on the tissue section, and is used to polarize the bounced target micro-region under the action of an electrostatic field, so that the target micro-region is adsorbed into the collector under the action of dielectrophoretic force.

[0006] Preferably, it further includes an electrostatic generator. An electrostatic generating head is connected between the electrostatic generator and the collector, and an electrostatic is uniformly applied to the collector.

[0007] Preferably, the region where laser scanning cutting is implemented is set in the region on the upper surface of the blade between the tip of the blade at the front end and 10 μm away from the tip.

[0008] Preferably, the beam expander is arranged between the laser and the galvanometer, and includes a first lens close to the laser and a second lens close to the galvanometer.

[0009] Preferably, the objective lens is located obliquely above the blade, and the optical axis of the focused light spot emitted by the objective lens is perpendicular to the upper surface of the blade.

[0010] Preferably, a plurality of collectors are arranged above the target micro-region in a vertical and horizontal array or honeycomb arrangement. The method for laser cutting and collecting the target micro-region of the tissue section performed on the blade includes the following steps: S1. Place the sample tissue block at the operation position on the carrier stage, image the sample tissue block, select the target micro-region to be cut, and store the tissue section position of the sample tissue block and the target position information of all target micro-regions; S2. Connect the collector to the electrostatic generator and adjust the position of the collector to ensure that the collector is located above the target micro-region; S3. The laser provides pulsed laser, which is expanded by a beam expander and scanned by a galvanometer mirror, and then focused by an objective lens into a light spot with energy reaching the set cutting threshold, and waits for a trigger signal to emit light. S4. The control module signals to control the carrier stage to drive the sample tissue block to move horizontally, ensuring that the blade and the sample tissue block move relative to each other to achieve mechanical sectioning. When moving to the tissue section position stored in step S1, the control module sends signals to the galvanometer mirror and the pulsed laser. By adjusting the scanning angle θ and scanning speed v of the galvanometer mirror, a series of target position points on the target micro-region are scanned and ablated one by one. When the last point of a closed curve is ablated, the shock wave pressure generated synchronously reflects on the blade surface to make the target micro-region bounce up. S5. Turn on the collector, and polarization occurs in the non-uniform electric field generated by the collector, so that the target micro-region is adsorbed into the collector under the action of dielectrophoretic force.

[0011] Preferably, in step S4, the galvanometer mirror adopts a single-axis galvanometer mirror horizontal scanning mode, and the scanning speed is controlled by setting the scanning time, thereby adjusting the cutting speed v of the pulsed laser in the feed direction to ensure that the interval between adjacent ablation points is less than the ablation point diameter.

[0012] Preferably, when the diameter of the target micro-region is less than 100 μm, the cutting and ejection of the target micro-region are realized by a focused light spot perpendicular to the upper surface of the blade. When the diameter of the target micro-region is greater than 100 μm, a horizontal focused beam is added that is emitted in the horizontal direction to increase the ejection force applied to the target micro-region.

[0013] Preferably, when there are multiple target micro-regions and multiple collectors are arranged in an array on the target micro-region, step S5 further includes, after each collector completes collection, sequentially adjusting the position of the empty collector above the next target micro-region for collection until there is no empty collector.

[0014] The beneficial effects of the technical solution of the present invention are mainly reflected in: 1. Combining laser microdissection with mechanical sectioning ensures the integration of the entire acquisition process of the sample's morphology from the tissue block to the tissue section and then to single cells or cell clusters (target micro-regions) in the tissue section, more precisely retaining the three-dimensional position information of the target cells or cell clusters, which is more conducive to realizing the spatial histology analysis of specific cells. At the same time, compared with the traditional laser microdissection method, there is no need for additional tissue sectioning and pasting steps. By tilting the blade for cutting and using the blade as the base of the tissue section, and directly performing laser scanning cutting on the tissue section on the blade surface, not only the cutting efficiency is improved, but also non-contact cutting can be achieved to prevent sample contamination.

[0015] 2. By utilizing the reflective ejection force generated during the cutting of tissue sections by focused laser and combining it with the dielectrophoretic force generated in the non-uniform electric field of a charged collector on the target micro-region, non-contact and rapid collection of the target micro-region placed on the inclined blade surface is achieved. Meanwhile, when there are multiple target micro-regions, by arranging multiple collectors in a vertical and horizontal array or a honeycomb pattern above the target micro-regions to collect different target micro-regions in sequence, the collection efficiency is further improved.

[0016] 3. Different laser cutting methods are adopted according to the different diameters of the target micro-regions, which has higher compatibility for the collection of target micro-regions of different sizes. For target micro-regions with a smaller diameter, the cutting and ejection of the target micro-region can be achieved through a focused light spot perpendicular to the upper surface of the blade. For target micro-regions with a larger diameter, a focused light beam emitted in the horizontal direction is added on the basis of the original focused light spot to increase the ejection force, facilitating the collection of large-diameter target micro-regions.

[0017] 4. By adjusting the scanning angle and scanning speed of the galvanometer scanner, the ablation point position and ablation point spacing of the focused light spot on the section are regulated. Meanwhile, by setting the scanning speed, the interval between adjacent ablation points is ensured to be less than the ablation point diameter, thereby ensuring the continuity of cutting and the complete cutting of the target micro-region. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 is a schematic structural diagram of a device for laser cutting and collection of target micro-regions of tissue sections on a blade; Figure 2 is a schematic principle diagram of a method for laser cutting and collection of target micro-regions of tissue sections on a blade; Figure 3 is a flowchart of a method for laser cutting and collection of target micro-regions of tissue sections on a blade; Figure 4 is a schematic principle diagram of laser cutting of a target micro-region by synchronously combining the scanning angle and scanning speed; Figure 5 is a schematic diagram of a method for cutting and collecting a target micro-region with a larger diameter; Figure 6 is a schematic diagram of a method for collecting a target micro-region in an embodiment with multiple collectors. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0019] To clearly and detailedly show the objectives, advantages, and features of the present invention, it will be illustrated and explained through non-restrictive descriptions of the following preferred embodiments. This embodiment is only a typical example of applying the technical solution of the present invention, and all technical solutions formed by equivalent replacement or equivalent transformation fall within the scope of protection required by the present invention.

[0020] At the same time, it is stated that in the description of the solution, it should be noted that the orientation or positional relationship indicated by terms such as "upper", "lower", "inner", "outer", etc. is based on the orientation or positional relationship shown in the drawings. It is only for the convenience of description and simplification, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation. Therefore, it should not be construed as a limitation to the present invention.

[0021] In addition, the terms "first" and "second" in this solution are only for descriptive purposes and cannot be construed as indicating or implying a ranking of importance or implicitly specifying the quantity of the indicated technical features. Therefore, the features defined with "first" and "second" may explicitly or implicitly include one or more of such features. In the present invention, the meaning of "a plurality" is two or more, unless otherwise specifically defined.

[0022] The present invention discloses a device for laser cutting and collecting target micro-regions of tissue sections performed on a blade 110. First, a tissue section needs to be made on a sample tissue block 107, and then the target micro-region 109 is separated by laser cutting on the tissue section 108. The target micro-region 109 can be a small-sized target micro-region 109, such as a single cell, or a large-sized target micro-region 109, such as a cell population. Such as Figure 1 、 Figure 2 As shown, the device includes: A carrier stage for placing the sample tissue block 107 and controlling its movement. During the entire cutting process, it is necessary to ensure that the sample tissue block 107 is always fixed at the operating position on the carrier stage.

[0023] A blade 110 is inclinedly arranged on one side of the carrier stage through a fixture. The blade 110 is stationary, and the carrier stage is moved to make the blade 110 move relative to the sample tissue block 107 on the carrier stage to complete mechanical sectioning and obtain a tissue section 108. After cutting, the tissue section 108 is located on the upper surface of the blade 110, and the blade 110 serves as the base of the tissue section 108. The inclination angle of the blade 110 is φ, so the inclination angle of the tissue section 108 located on the upper surface of the blade 110 is also φ.

[0024] In some embodiments, the material of the blade 110 can be selected from one of diamond, glass knife, steel knife, and tungsten carbide knife.

[0025] In some embodiments, the position and angle of the blade 110 on the fixture are adjustable. Before mechanical slicing, the position and angle of the blade 110 on the fixture are precisely adjusted first, so as to control the uniformity of the overall thickness of the tissue slice 108 during the movement and cutting process of the carrier stage. In some embodiments, the inclination angle φ is 30°-45°. When the inclination angle φ of the blade 110 is within this range, the overall thickness of the slice is relatively uniform.

[0026] A laser cutting system includes a laser 101 for providing pulsed laser, a beam expander for expanding the laser emitted by the laser 101, a galvanometer scanner 104 for controlling the scanning angle and scanning speed of the expanded laser, and an objective lens 106 for focusing the scanned laser beam. In one embodiment, a reflecting mirror 105 for reflecting the beam scanned by the galvanometer scanner 104 to the objective lens 106 is further disposed between the galvanometer scanner 104 and the objective lens 106; wherein, the objective lens 106 focuses the scanned laser into a light spot reaching the cutting threshold, and uses this light spot to complete the cutting of the target micro-region 109. By adjusting the scanning angle θ of the galvanometer scanner 104 and combining the focusing of the scanned laser by the objective lens 106, the cutting of the tissue slice 108 and the target micro-region 109 on the tissue slice 108 and the bouncing of the target micro-region 109 are achieved.

[0027] A control module 114 is respectively connected to the carrier stage and the laser cutting system in a signal connection, and synchronously controls mechanical slicing and laser cutting. Among them, the control module 114 is at least connected to the carrier stage, the laser 101 and the galvanometer scanner 104 in a signal connection. During the working process, the control module 114 real-time regulates the moving position of the carrier stage, and synchronously triggers the laser emission of the laser 101, the scanning angle and scanning speed of the galvanometer scanner 104, etc. in the laser cutting system, so as to achieve the cutting and capture of the target micro-region 109.

[0028] A collector 111 is located above the target micro-region 109 on the tissue slice 108, and is used to polarize the bounced target micro-region 109 under the action of an electrostatic field, so that the target micro-region 109 is adsorbed from the bottom of the collector 111 into the collector 111 under the action of dielectrophoresis force.

[0029] In some embodiments, an electrostatic generator 113 is further provided. An electrostatic generating head 112 is connected between the electrostatic generator 113 and the collector 111. Among them, the electrostatic generator 113 is used to generate static electricity, and the electrostatic generating head 112 is used to uniformly apply static electricity to the collector 111, so as to ensure that the collector 111 is charged with static electricity, and the bounced target micro-region 109 is polarized under the action of the electrostatic field, and thus is adsorbed and collected under the action of dielectrophoresis force. Among them, the optimal distance between the collector 111 and the electrostatic generating head 112 can also be measured through experiments, and the collector 111 and the electrostatic generating head 112 are adjusted to the optimal distance through a fixing device, so that the collector 111 can store more static electricity, thereby generating a greater electric field strength, improving the collection efficiency and collection stability. In some embodiments, the collector 111 can be made of a material with a relatively large dielectric constant, so as to facilitate storing more static electricity, generating a stronger electric field, making the polarization intensity of the target micro-region 109 greater, and the dielectrophoresis adsorption force stronger.

[0030] Among them, after the mechanical sectioning is completed, the blade 110 at the bottom of the tissue section 108 can be used as a glass slide for carrying the tissue section 108. To ensure the quality of laser cutting, the tissue section 108 needs to be in good contact with the upper surface of the blade 110. The area where laser scanning cutting is performed is set in the area between the tip of the blade at the front end of the blade and 10 μm away from the tip. Since the tip of the front end of the blade first contacts and cuts the sample tissue block 107, and the blade is inclined, after the cutting is completed, the closer the position of the blade to the tip of its front end is to the bottom of the tissue section 108, the better the degree of fit. Therefore, using the area between the tip of the front end of the blade and 10 μm away from the tip as the area for performing laser scanning cutting can better ensure the quality of laser cutting.

[0031] In some embodiments, the laser 101 is an ultraviolet pulsed laser 101. The ultraviolet pulsed laser 101 has a shorter wavelength, which is convenient for generating a smaller focused spot. Under the condition of constant pulse energy, it can achieve a higher energy density, and at the same time, it can improve the precision of laser cutting.

[0032] In some other embodiments, the laser 101 can also be a picosecond pulsed laser 101. Compared with nanosecond pulses, picosecond pulses have non-linear absorption and can generate a strong shock wave pressure within a short distance with a smaller pulse energy. Thus, while achieving cutting, the generated shock wave is sufficient to cause the target micro-region 109 to bounce at the position of the last point where it is laser-segmented, thereby helping the subsequent collection of the target micro-region 109.

[0033] The beam expander is disposed between the laser 101 and the galvanometer 104, and includes a first lens 102 close to the laser 101 and a second lens 103 close to the galvanometer 104. In some embodiments, the first lens 102 and the second lens 103 can be made of high-purity fused silica material and coated with an ultraviolet antireflection film to ensure high transmittance and reduce energy loss. The beam expansion ratio of the beam expander can be adjusted in real time as needed, which will not be elaborated here.

[0034] The objective lens 106 is located obliquely above the blade 110. In a preferred embodiment, the optical axis of the focused light spot emitted by the objective lens 106 is perpendicular to the upper surface of the blade 110 to ensure the accuracy of laser cutting.

[0035] A plurality of collectors 111 can be arranged on the target micro-region 109 to improve the collection efficiency and throughput. In some embodiments, a plurality of collectors 111 are arranged in a vertical and horizontal array above the target micro-region 109. The number of collectors 111 arranged in the vertical and horizontal columns, for example: M×N (1≤M≤10, 1≤N≤10, M and N are both positive integers). During the collection process, a group of target micro-regions 109 can be continuously cut on the section by laser cutting in advance, and then a plurality of collectors 111 above a group of target micro-regions 109 are synchronously turned on for collection. After the collection is completed, the position of the next group of collectors 111 is adjusted, and the next group of target micro-regions 109 are cut and collected; in other embodiments, a plurality of collectors 111 can also be arranged in a honeycomb shape above the target micro-region 109.

[0036] In some embodiments, a variety of imaging modules can also be provided that are coupled to the target micro-region laser cutting and collection device for tissue sectioning on the blade 110. The imaging module is used for imaging the surface of the sample tissue block 107 and positioning the cutting position of the target micro-region 109. Before the carrier stage drives the sample tissue block 107 to move to the bottom of the blade 110, it first moves to the imaging module for imaging, selects the target micro-region 109 to be cut, and records the position information to facilitate subsequent synchronous mechanical cutting and laser cutting.

[0037] As Figure 3 shown, the method for laser cutting and collecting the target micro-region 109 of the tissue section 108 on the blade 110 includes the following steps: S1. Place the sample tissue block 107 at the operation position on the carrier stage, image the sample tissue block 107, select the target micro-region 109 to be cut, and store the tissue section 108 position of the sample tissue block 107 and the target position information of all target micro-regions 109.

[0038] S2. The collector 111 is connected to the electrostatic generator 113, and the position of the collector 111 is adjusted to ensure that the collector 111 is located above the target micro-region 109.

[0039] S3. The laser 101 provides pulsed laser light. After being expanded by the beam expander and scanned by the galvanometer 104, the pulsed laser light is focused by the objective lens 106 into a light spot with an energy reaching the set cutting threshold, and waits for the trigger signal to emit light.

[0040] S4. The control module 114 controls the signal to drive the sample tissue block 107 to move horizontally, ensuring that the blade 110 and the sample tissue block 107 move relative to each other to achieve mechanical sectioning. When moving to the position of the tissue section 108 stored in step S1, the control module 114 sends signals to the galvanometer 104 and the pulsed laser. By adjusting the scanning angle θ and scanning speed v of the galvanometer 104, a series of target position points on the target micro-region 109 are scanned and ablated one by one. When the last point of a closed curve is ablated, the shock wave pressure generated synchronously is reflected on the knife surface to make the target micro-region 109 bounce up.

[0041] In some embodiments, in step S4, the galvanometer 104 adopts a single-axis galvanometer 104 horizontal scanning mode. The ablation point position of the pulsed laser is controlled by adjusting the scanning angle θ of the galvanometer 104, and the scanning speed is controlled by setting the scanning time, thereby adjusting the cutting speed v of the pulsed laser in the feed direction to ensure that the interval between adjacent ablation points is less than the ablation point diameter.

[0042] As Figure 4 shown, taking the upper left corner of the sample tissue block 107 as the coordinate origin o to establish a coordinate system, imaging its surface before sectioning, storing the position information of the target micro-region 109. After that, the sample tissue block 107 is cut into a tissue section 108 by the blade 110 and generates an inclination angle φ with the blade 110. At this time, taking the upper left corner of the tissue section 108 as the coordinate system origin O', and performing laser cutting on the target micro-region 109 on the tissue section 108. To reduce the complexity of the transformation of the target position information, the single-axis galvanometer 104 horizontal scanning mode is adopted, and only the scanning angle along the x direction needs to be set, as Figure 4As shown, for two ablation points at the same height in the x direction, horizontal scanning is performed, scanning from ablation point 1 to 2, and then from ablation point 3 to 4, so as to accurately determine the target positions of each ablation point for cutting on the target micro-region 109. For the y direction, that is, the feed direction, the scanning speed of the galvanometer 104 is determined by controlling the scanning time, and then the cutting speed v is accurately controlled to ensure that the interval Δn between two adjacent ablation points at different heights (such as ablation point 1 and ablation point 3) is less than the diameter of the ablation point, avoiding the existence of uncut parts between adjacent ablation points, so as to achieve cutting continuity and ensure the complete separation of the target micro-region 109; in addition, considering that the fitting area between the tissue section 108 and the surface of the blade 110 is extremely small, this horizontal scanning combined with mechanical cutting method can not only achieve precise synchronization, but also make the laser beam for ablation always located at the focal plane of the target micro-region 109, ensuring the cutting quality.

[0043] S5. Turn on the collector 111, and polarization occurs in the non-uniform electric field generated by the collector 111, so that the target micro-region 109 is adsorbed into the collector 111 under the action of dielectrophoretic force.

[0044] Among them, when the diameter of the target micro-region 109 is less than 100 μm, at this time, the diameter of the target micro-region 109 is small and the area gravity is also small, and the cutting and ejection of the target micro-region 109 can be realized through the focused light spot in the direction perpendicular to the upper surface of the blade 110; As Figure 5 shown, when the diameter of the target micro-region 109 is greater than 100 μm, since the diameter of the target micro-region 109 is large at this time, the area and gravity of the target micro-region 109 also increase accordingly. At this time, the shock wave generated by the focused laser beam at the edge of the target micro-region 109 is not sufficient to cause the ejection of the target micro-region 109, nor can it reach the range of the electrostatic field generated by the upper collector 111. Therefore, the focused light spot in the vertical direction cannot realize the cutting and ejection of the target micro-region 109. To ensure the smooth collection of the target micro-region 109, a horizontal focused beam is added that is emitted in the horizontal direction to increase the ejection force applied to the target micro-region 109 to improve the ejection force. The shock wave pressure in the direction perpendicular to the beam generated when interacting with the tissue is used to lift the target micro-region 109 with a larger diameter. At this time, due to the large area of the target micro-region 109, the local damage of the horizontal laser beam 116 to the target micro-region 109 in the horizontal direction can be ignored.

[0045] As Figure 6As shown, when there are multiple target micro-regions 109 and multiple collectors 111 are arranged in an array on the target micro-regions 109, step S5 further includes, after each collector 111 finishes collection, sequentially adjusting the positions of the remaining collectors 111 above the next target micro-region 109 for collection until there are no remaining collectors 111. Among them, after laser cutting a row of target micro-regions 109, the collectors 111 in the same row sequentially collect the corresponding target micro-regions 109 (as Figure 6 shown, sequentially collecting 109a, 109b, 109c, etc.), and gradually adjusting the positions of the moving collectors 111 during the collection process; after the collection of the target micro-regions 109 in the same row is completed, the next row of target micro-regions 109 is cut and collected. Before collecting the next row of target micro-regions 109, the corresponding row of collectors 111 is moved above the target micro-regions 109 to be collected; Figure 6 In Figure 6 , a standard eight-connected tube collector 117 is used to be compatible with a centrifuge and facilitate subsequent RNA and protein sequencing.

[0046] There are still various implementation manners of the present invention. All technical solutions formed by using equivalent transformations or equivalent substitutions fall within the protection scope of the present invention.

Claims

1. A target micro-region laser cutting and collecting device for tissue sections performed on a blade, characterized in that: Comprising: A stage for placing a sample tissue block and controlling its movement; A blade, which is obliquely arranged on one side of the stage through a fixture. The inclination angle of the blade is φ. The blade is stationary, and the stage is moved to make the blade move relative to the sample tissue block on the stage to complete mechanical sectioning and obtain tissue sections. After cutting, the tissue sections are located on the upper surface of the blade, and the blade serves as the base of the tissue sections; A laser cutting system, including a laser for providing pulsed laser, a beam expander for expanding the laser emitted by the laser, a galvanometer for controlling the scanning angle and scanning speed of the expanded laser, and an objective lens for focusing the scanned laser beam. By adjusting the scanning angle θ of the galvanometer and combining with the objective lens, the scanned laser is focused to achieve cutting of the tissue sections and the target micro-regions on the tissue sections and the bouncing of the target micro-regions; A control module, which is signal-connected to the stage and the laser cutting system respectively, and synchronously controls mechanical sectioning and laser cutting; A collector, which is located above the target micro-region on the tissue section, and is used to polarize the bounced target micro-region under the action of an electrostatic field, so that the target micro-region is adsorbed into the collector under the action of dielectrophoresis force.

2. The target micro-region laser cutting and collection device for tissue sections performed on a blade according to claim 1, characterized in that: It further includes an electrostatic generator, and an electrostatic generating head is connected between the electrostatic generator and the collector, and an electrostatic is uniformly applied to the collector.

3. The laser cutting and collection device for the target micro-region of tissue sections performed on the blade according to claim 1, characterized in that: The area where laser scanning cutting is implemented is set in the area on the upper surface of the blade between the tip of the blade at the front end of the blade and 10 μm away from the tip.

4. The laser cutting and collection device for the target micro-region of tissue sections performed on a blade according to claim 1, characterized in that: The beam expander is arranged between the laser and the galvanometer, and includes a first lens close to the laser and a second lens close to the galvanometer.

5. The laser cutting and collection device for the target micro-region of tissue sections performed on the blade according to claim 1, characterized in that: The objective lens is located obliquely above the blade, and the optical axis of the focused light spot emitted by the objective lens is perpendicular to the upper surface of the blade.

6. The target micro-region laser cutting and collection device for tissue sections performed on a blade according to claim 1, characterized in that: Above the target micro-region, a plurality of collectors are arranged in a horizontal and vertical array or in a honeycomb pattern.

7. A method for laser cutting and collecting target micro-regions of tissue sections performed on a blade, characterized in that: Including the following steps: S1. Place the sample tissue block at the operation position on the stage, image the sample tissue block, select the target micro-region to be cut, and store the tissue section position of the sample tissue block and the target position information of all target micro-regions; S2. Connect the collector to the electrostatic generator, and adjust the position of the collector to ensure that the collector is located above the target micro-region; S3. The laser provides pulsed laser, which is expanded by the beam expander and scanned by the galvanometer, and then focused by the objective lens into a light spot with energy reaching the set cutting threshold, and waits for the trigger signal to emit light; S4. The control module signals to control the stage to drive the sample tissue block to move horizontally, ensuring that the blade moves relative to the sample tissue block to achieve mechanical sectioning. When moving to the tissue section position stored in step S1, the control module sends signals to the galvanometer and the pulsed laser. By adjusting the scanning angle θ and scanning speed v of the galvanometer, a series of target position points on the target micro-region are scanned and ablated one by one. When the last point of a closed curve is ablated, the shock wave pressure generated synchronously is reflected on the blade surface to make the target micro-region bounce; S5. Turn on the collector, polarize in the non-uniform electric field generated by the collector, and make the target micro-region be adsorbed into the collector under the action of dielectrophoresis force.

8. The method for laser cutting and collecting the target micro-region of the tissue section performed on the blade according to claim 7, characterized in that: In step S4, the galvanometer uses a single-axis galvanometer for lateral scanning. The scanning speed is controlled by setting the scanning time, and then the cutting speed v of the pulsed laser in the feed direction is adjusted to ensure that the interval between adjacent ablation points is less than the ablation point diameter.

9. The method for laser cutting and collecting of a target micro-region on a blade according to claim 7, characterized in that: When the diameter of the target micro-region is less than 100 μm, the cutting and ejection of the target micro-region are achieved by a focused light spot perpendicular to the upper surface of the blade. When the diameter of the target micro-region is greater than 100 μm, a horizontal focused beam is added that is emitted in the horizontal direction to increase the ejection force applied to the target micro-region.

10. The method for laser cutting and collecting the target micro-region of the tissue section performed on the blade according to any one of claims 7-9, characterized in that: When there are multiple target micro-regions and multiple collectors are arranged in an array on the target micro-region, step S5 further includes, after each collector completes collection, successively adjusting the position of the vacant collector above the next target micro-region for collection until there is no vacant collector.

Images