Biological three-dimensional microscopic imaging system

Abstract

The utility model relates to the technical field of biological imaging, and provides a biological three-dimensional microscopic imaging system which comprises a sample bearing module which is suitable for placing a biological sample, is used for alternately driving the biological sample to move between a milling station and an imaging station, and drives the biological sample to be lifted by a preset height after moving to the milling station; the sample plane processing module is arranged at the milling station; the scanning module is arranged at the imaging station; and the control processing module is used for acquiring the plurality of cross-section images according to the scanning module so as to obtain a three-dimensional image of the biological sample according to the plurality of cross-section images of the sequence. The biological three-dimensional microscopic imaging system provided by the utility model has the advantages that the automation degree of the system is high, and the sample bearing module, the sample plane processing module and the scanning module work cooperatively, so that high-speed, high-smoothness, traceless and high-precision surface removal can be carried out on a biological sample, the rapid and continuous treatment of the biological sample is realized, and the imaging efficiency is improved.

Description

TECHNICAL FIELD

[0001] The utility model relates to biological imaging technical field especially relates to a biological three -dimensional microscopic imaging system. BACKGROUND

[0002] Traditional microscope imaging technology is mainly limited to two-dimensional space, through the slice of biological body or other observation object, and selects the fragment among them to carry out two-dimensional imaging analysis. However, biological body and many other substances essentially have three-dimensional structure, and only two-dimensional imaging information is used to deduce its three-dimensional characteristics, which often leads to serious information loss, and even can cause misjudgment. Therefore, in recent years, three-dimensional microscopic imaging technology has become the hotspot and development direction in biomedical research field.

[0003] Among numerous three-dimensional microscopic imaging technologies, fMOST (fluorescence Micro-Optical Sectioning Tomography) technology and light sheet technology (Light Sheet Fluorescence Microscopy, LSFM) are particularly attractive.

[0004] fMOST technology is a kind of three-dimensional microscopic imaging technology combining physical sectioning and fluorescence imaging. In this technology, the fixed sample is continuously sliced layer by layer, and the thickness of each layer can reach microns. After slicing, the sample surface remains flat, and no additional polishing treatment is required to directly scan and image by fluorescence microscope. This process is repeated until the information of each layer of biological tissue is obtained. Then, the information is reconstructed by software algorithm, so as to obtain the three-dimensional spatial structure of the biological sample. However, fMOST technology faces a significant pain point in practical application: diamond slicing knife is easy to produce regenerative chatter during continuous cutting, that is, the ripples left by the previous cutting will periodically stimulate the tool, aggravate the vibration amplitude, and result in slow cutting speed.

[0005] On the other hand, light sheet technology is a kind of three-dimensional microscopic imaging technology that uses a thin layer of laser to illuminate the sample from the side (perpendicular to the detection axis). This technology only excites fluorescence signals in the focal plane, and completes three-dimensional imaging by moving the sample or light sheet. Light sheet technology has the advantages of fast imaging speed and low phototoxicity, and is particularly suitable for long-term dynamic observation of living cells. However, light sheet technology also has its limitations. Due to the limited imaging distance of the objective lens, this technology cannot image samples with large height, which limits its application in large biological samples or tissue block research. UTILITY MODEL CONTENT

[0006] The utility model provides a kind of biological three-dimensional microscopic imaging system to solve the problem of slow cutting speed of existing imaging technology, which cannot image samples with large height.

[0007] The utility model provides a kind of biological three-dimensional microscopic imaging system, comprising:

[0008] Sample bearing module is suitable for arranging biological sample, for alternately driving the biological sample moves between milling station and imaging station, and after moving to the milling station, the biological sample is driven to lift preset height;

[0009] Sample plane processing module is set in milling station, including drive assembly and milling cutter, the drive end of the drive assembly is connected with the milling cutter, the drive assembly is used to cooperate the movement of the sample bearing module to mill the biological sample when the biological sample moves to the milling station, to form smooth surface without trace on the section of the biological sample;

[0010] Scanning module is set in imaging station, for cooperating the movement of the sample bearing module to image the smooth surface without trace of the biological sample when the biological sample moves to the imaging station, to obtain section image;

[0011] Control processing module is electrically connected with the scanning module, for obtaining a plurality of section images according to the scanning module, to obtain three-dimensional image of the biological sample according to a plurality of sequence section images.

[0012] According to the biological three-dimensional microscopic imaging system provided by the utility model, the drive assembly comprises:

[0013] Air compression motor;

[0014] Pneumatic spindle, which is provided with a first mounting cavity matched with the milling cutter in axial direction, and is driven to rotate at high speed by air pressure of the air compression motor;

[0015] First clamping piece is arranged at the opening end of the first mounting cavity, with adjustable first clamping structure, for coaxially locking milling cutter of different diameter specifications on the rotation center axis of the pneumatic spindle.

[0016] According to the biological three-dimensional microscopic imaging system provided by the utility model, the drive assembly comprises:

[0017] Drive motor;

[0018] Electric spindle, which is connected with the output shaft of the drive motor through coupling, and is provided with a second mounting cavity matched with the milling cutter in axial direction;

[0019] Second clamping piece is arranged at the opening end of the second mounting cavity, with adjustable second clamping structure, for coaxially locking milling cutter of different diameter specifications on the rotation center axis of the electric spindle.

[0020] According to the biological three-dimensional microscopic imaging system, the milling cutter comprises a handle and a cutter head, the diameter of the handle is 2-6mm, and the cutter head is a diamond cutter head.

[0021] According to the biological three-dimensional microscopic imaging system, the sample bearing module is a three-dimensional precision platform, and the three-dimensional precision platform comprises:

[0022] The X-axis moving platform, the Y-axis moving platform and the Z-axis moving platform are sequentially stacked from bottom to top, the biological sample is arranged on the Z-axis moving platform, the Z-axis moving platform is used for driving the biological sample to move along the height direction of the Z-axis moving platform, the Y-axis moving platform is used for driving the Z-axis moving platform and the biological sample to move along the width direction of the biological sample, and the X-axis moving platform is used for driving the Y-axis moving platform, the Z-axis moving platform and the biological sample to move along the length direction of the biological sample.

[0023] Alternatively, the Z-axis moving platform, the X-axis moving platform and the Y-axis moving platform are sequentially stacked from bottom to top, the biological sample is arranged on the Y-axis moving platform, the Y-axis moving platform is used for driving the biological sample to move along the width direction of the Y-axis moving platform, the X-axis moving platform is used for driving the Y-axis moving platform and the biological sample to move along the length direction of the biological sample, and the Z-axis moving platform is used for driving the X-axis moving platform, the Y-axis moving platform and the biological sample to move along the height direction of the biological sample.

[0024] According to the biological three-dimensional microscopic imaging system, the scanning module is a TDI line scanning imaging device, which comprises a TDI camera, a first illumination mechanism, a first detection mechanism and a first objective lens.

[0025] The first illumination mechanism irradiates the section of the biological sample through the first objective lens, and the TDI camera and the first detection mechanism shoot the section of the biological sample through the first objective lens.

[0026] According to the biological three-dimensional microscopic imaging system, the scanning module is a wide-field surface imaging device, which comprises a surface array camera, a second illumination mechanism, a second detection mechanism and a second objective lens.

[0027] The second illumination mechanism irradiates the biological sample through the second objective lens, and the surface array camera and the second detection mechanism shoot the section of the biological sample through the second objective lens.

[0028] The utility model provides a kind of biological three-dimensional microscopic imaging system, the scanning module is oblique light sheet imaging device, it include: third illumination mechanism, third detection mechanism, third objective and fourth objective;

[0029] The third illumination mechanism is obliquely irradiated on one side of the section of the biological sample by the third objective, and the third detection mechanism is obliquely photographed on the other side of the section of the biological sample by the fourth objective.

[0030] The utility model provides a kind of biological three-dimensional microscopic imaging system, the scanning module is vertical light sheet imaging device, it include: fourth illumination mechanism, fifth illumination mechanism, fourth detection mechanism, fifth objective, sixth objective and seventh objective;

[0031] The fourth illumination mechanism is vertically irradiated on one side of the biological sample by the fifth objective, the fifth illumination mechanism is vertically irradiated on the other side of the biological sample by the sixth objective, and the fourth detection mechanism is vertically photographed on the section of the biological sample by the seventh objective.

[0032] The biological three-dimensional microscopic imaging system provided by the utility model accurately controls the alternate movement of biological sample between milling station and imaging station by sample bearing module, and the lifting of sample surface after milling, to ensure the continuity and accuracy of imaging process. The milling cutter in sample plane processing module can accurately mill biological sample under the driving of driving assembly, to form a smooth surface without trace, to provide high-quality section for subsequent scanning imaging. The scanning module performs high-resolution imaging on the smooth surface without trace, and the control processing module processes multiple section images, to reconstruct high-precision three-dimensional image of biological sample. The system has high automation degree, and the cooperative work between sample bearing module, sample plane processing module and scanning module can perform high-speed, high-smoothness, traceless and high-precision surface removal on biological sample, to realize rapid and continuous processing of biological sample, and improve imaging efficiency. BRIEF DESCRIPTION OF DRAWINGS

[0033] In order to more clearly illustrate the technical scheme of the utility model or prior art, the following will be a brief introduction to the drawings needed to be used in the embodiment or prior art description, obviously, the drawings in the following description are some embodiments of the utility model, and for those skilled in the art, other drawings can be obtained without creative labor according to these drawings.

[0034] Figure 1 It is one of the schematic diagrams of the biological three-dimensional microscopic imaging system provided by the utility model.

[0035] Figure 2It is the schematic view two of biological three-dimensional microscopic imaging system provided by the utility model.

[0036] Figure 3 It is the schematic view two of biological three-dimensional microscopic imaging system provided by the utility model.

[0037] Figure 4 It is the schematic view one of scanning module provided by the utility model.

[0038] Figure 5 It is the schematic view two of scanning module provided by the utility model.

[0039] Figure 6 It is the schematic view three of scanning module provided by the utility model.

[0040] Figure 7 It is the schematic view four of scanning module provided by the utility model.

[0041] Figure 8 It is the flow schematic view of control method of biological three-dimensional microscopic imaging system provided by the utility model.

[0042] Reference signs:

[0043] 1, sample bearing module; 11, X-axis moving platform; 12, Y-axis moving platform; 13, Z-axis moving platform;

[0044] 2, sample plane processing module; 21, milling cutter; 22, pneumatic spindle; 23, first clamping piece; 24, electric spindle; 25, second clamping piece;

[0045] 3, scanning module; 30, fourth illumination mechanism; 31, TDI camera; 32, first illumination mechanism; 33, first detection mechanism; 34, area array camera; 35, second illumination mechanism; 36, second detection mechanism; 37, third illumination mechanism; 38, third detection mechanism; 39, fifth illumination mechanism; 391, fourth detection mechanism;

[0046] 4, control processing module;

[0047] 5, biological sample. DETAILED DESCRIPTION

[0048] In order to make the purpose, technical scheme and advantages of the utility model more clear, the technical scheme in the utility model will be described clearly and completely below in combination with the drawings in the utility model. Obviously, the described embodiments are a part of the embodiments of the utility model, rather than all the embodiments. Based on the embodiments in the utility model, all other embodiments obtained by the person skilled in the art without creative labor belong to the protection scope of the utility model.

[0049] The biological three-dimensional microscopic imaging system and the control method thereof are described below. Figures 1 to 8 The biological three-dimensional microscopic imaging system and the control method thereof are described below.

[0050] The biological three-dimensional microscopic imaging system and the control method thereof are described below. Figure 1 The biological three-dimensional microscopic imaging system and the control method thereof are described below. Figure 3 As shown in the figure, the biological three-dimensional microscopic imaging system comprises a sample bearing module 1, a sample plane processing module 2, a scanning module 3 and a control processing module 4. The sample bearing module 1 is suitable for placing a biological sample 5, and is used for alternately driving the biological sample 5 to move between a milling station and an imaging station, and driving the biological sample 5 to be lifted by a preset height after moving to the milling station; the sample plane processing module 2 is arranged at the milling station and comprises a driving assembly and a milling cutter 21, the driving end of the driving assembly is connected with the milling cutter 21, and the driving assembly is used for milling the biological sample 5 in cooperation with the movement of the sample bearing module 1 when the biological sample 5 moves to the milling station, so as to form a smooth and mark-free surface on the section of the biological sample 5; the scanning module 3 is arranged at the imaging station and is used for imaging the smooth and mark-free surface of the biological sample 5 in cooperation with the movement of the sample bearing module 1 when the biological sample 5 moves to the imaging station, so as to obtain a section image; the control processing module 4 is electrically connected with the scanning module 3 and is used for obtaining a plurality of section images according to the scanning module 3, so as to obtain a three-dimensional image of the biological sample 5 according to the sequence of the plurality of section images.

[0051] The imaging steps of the biological three-dimensional microscopic imaging system are as follows:

[0052] In this embodiment, the biological sample 5 is first prepared. The biological sample is gradient dehydrated until completely dehydrated. The penetration of resin monomer and crosslinking agent is performed. The penetrated resin is polymerized at low temperature / high temperature or ultraviolet, to form a solid block. The resin material can be selected from natural resin or synthetic resin (such as epoxy resin, polyester resin, polyurethane resin, phenolic resin and acrylic resin), and can be prepared by free radical polymerization, ionic polymerization or polycondensation reaction, to obtain the biological sample 5.

[0053] The prepared biological sample 5 is arranged on the sample bearing module 1. The sample bearing module 1 drives the biological sample 5 to move to the milling station, and the sample plane processing module 2 mills the surface of the biological sample 5 in cooperation with the sample bearing module 1, for example, the sample bearing module 1 moves along the X axis and the Y axis, and the sample plane processing module 2 mills, to remove the first layer of the biological sample 5 (usually about 0.1 mm thick), to realize the surface milling.

[0054] The sample carrying module 1 drives the biological sample 5 to move to the milling station and is lifted by a preset height (usually 1 micrometer) along the Z axis. The sample planar processing module 2 mills the biological sample 5 in cooperation with the sample carrying module 1, for example, the sample carrying module 1 moves along the X axis and the Y axis, and the sample planar processing module 2 mills to form a smooth section. The sample carrying module 1 drives the biological sample 5 to move to the imaging station. The scanning module 3 cooperates with the movement of the sample carrying module 1 to image the smooth surface of the biological sample 5, and if the biological sample 5 is large, the sample carrying module 1 can be controlled to move along the X axis and the Y axis, and the scanning module 3 can take pictures to obtain a section image.

[0055] Then the biological sample 5 is alternately controlled to mill and image, that is, the plane milled by the biological sample 5 is milled again and imaged to obtain another section image at a preset height of the bottom of the section. Until a sufficient number of section images are obtained or the entire biological sample 5 is milled, the control processing module 4 reconstructs the three-dimensional image of the biological sample 5 according to the corresponding sequence of multiple section images.

[0056] In this embodiment, the control processing module 4 includes a control storage unit and a data processing unit. The control storage unit stores instructions and programs required for system operation, which instruct how each module works cooperatively. The user can set imaging parameters such as scanning speed, milling depth, imaging resolution, etc. through the control storage unit to meet the imaging needs of different biological samples 5. The control storage unit can also monitor the running state of the system in real time, including the working state of each module, temperature, humidity and other environmental parameters, to ensure that the system is in the best working state.

[0057] The data processing unit is responsible for processing and analyzing the data generated during the imaging process. Its main functions include: receiving the section image data transmitted by the scanning module 3 and performing preliminary preprocessing such as denoising and contrast enhancement. Since the biological sample 5 may have a slight displacement or deformation during milling and imaging, the data processing unit needs to register these section images to ensure that they can be accurately spliced together. Based on the registered section images, the data processing unit generates a three-dimensional image of the biological sample 5 using a three-dimensional reconstruction algorithm. In addition to three-dimensional reconstruction, the data processing unit can further analyze the generated three-dimensional image, such as measuring size, calculating volume, and identifying shape, to meet the different needs of users.

[0058] The biological three-dimensional microscopic imaging system provided by the utility model, through the sample bearing module 1 accurate control biological sample 5 in the milling station and the imaging station between the alternate movement, and the sample surface after milling, ensure the continuity and accuracy of the imaging process. The milling cutter 21 in the sample plane processing module 2 is driven under the driving assembly, can accurately mill biological sample 5, form the surface of no trace, provide high quality cross section for subsequent scanning imaging. The scanning module 3 carries out high resolution imaging to the surface of no trace, combines the control processing module 4 and handles multiple cross section images, can reconstruct the high precision three-dimensional image of biological sample 5. The system degree of automation is high, the cooperative work between sample bearing module 1, sample plane processing module 2 and scanning module 3 can carry out high speed, high finish, traceless, high precision surface removal to biological sample 5, realizes the rapid, continuous processing of biological sample 5, improves the imaging efficiency.

[0059] In some embodiments, as shown in Figure 1 The driving assembly includes an air compression motor, a pneumatic spindle 22 and a first clamping piece 23. The pneumatic spindle 22 is provided with a first installation cavity matched with the milling cutter 21 in the axial direction, and the pneumatic spindle 22 is driven by the air pressure of the air compression motor to rotate at high speed. The first clamping piece 23 is arranged at the opening end of the first installation cavity and has an adjustable first clamping structure for coaxially locking the milling cutter 21 of different diameters on the rotation center axis of the pneumatic spindle 22.

[0060] In this embodiment, the air compression motor provides air pressure power. The air compression motor generates high-pressure gas by compressing air to provide a stable air pressure driving source for the pneumatic spindle 22. The pneumatic spindle 22 is driven by the air pressure of the air compression motor to rotate at high speed. The high-speed rotating spindle can drive the milling cutter 21 to mill the biological sample 5 at a sufficient speed and power to form a smooth and traceless cross section. The first clamping piece 23 is arranged at the opening end of the first installation cavity and has an adjustable first clamping structure. This clamping structure can be adjusted according to the diameter of the milling cutter 21 to ensure that the milling cutter 21 can be coaxially locked on the rotation center axis of the pneumatic spindle 22. The main function of the first clamping piece 23 is to ensure that the milling cutter 21 is stably and accurately fixed in the installation cavity to prevent loosening or deviation during high-speed rotation, thereby affecting the milling quality.

[0061] When the biological sample 5 is placed on the milling station, the control processing module 4 sends a command to the air compressor motor to start working and generate air pressure. This air pressure is transmitted to the pneumatic spindle 22 through a pipe, driving the spindle to rotate at high speed. At the same time, the first clamping member 23 has adjusted the clamping structure according to the diameter of the milling cutter 21 and coaxially locks the milling cutter 21 onto the pneumatic spindle 22. With the cooperation of the sample carrying module 1, the biological sample 5 will gradually move to the milling position, at which point the milling cutter 21 driven by the pneumatic spindle 22 will begin to mill the sample.

[0062] In some embodiments, such as Figure 2 As shown, the drive assembly includes a drive motor, an electric spindle 24, and a second clamping member 25. The electric spindle 24 is connected to the output shaft of the drive motor via a coupling, and has a second mounting cavity adapted to the milling cutter 21 in the axial direction. The second clamping member 25 is disposed at the opening end of the second mounting cavity and has an adjustable second clamping structure for coaxially locking milling cutters 21 of different diameters onto the rotation center axis of the electric spindle 24.

[0063] In this embodiment, a drive motor provides rotational power. The drive motor transmits torque through its output shaft, driving the electric spindle 24 to rotate. The electric spindle 24 has a second mounting cavity adapted to the milling cutter 21. The electric spindle 24 is connected to the output shaft of the drive motor via a coupling, achieving stable power transmission. Driven by the drive motor, the electric spindle 24 achieves high-speed rotation. A second clamping member 25 is disposed at the opening end of the second mounting cavity and has an adjustable second clamping structure. The second clamping structure can be adjusted according to different milling cutter 21 diameters, ensuring that the milling cutter 21 can be coaxially locked on the rotation center axis of the electric spindle 24. The main function of the second clamping member 25 is to ensure that the milling cutter 21 is stably and accurately fixed within the mounting cavity, preventing loosening or displacement during high-speed rotation, thereby affecting milling quality and safety.

[0064] When the biological sample 5 is placed on the milling station, the control processing module 4 sends a command to the drive motor to start working and generate rotational power. This power is transmitted to the electric spindle 24 through a coupling, driving the spindle to rotate at high speed. At the same time, the second clamping member 25 has been adjusted according to the diameter of the selected milling cutter 21 and coaxially locks the milling cutter 21 onto the electric spindle 24. With the cooperation of the sample carrying module 1, the biological sample 5 will gradually move to the milling position, at which point the milling cutter 21 driven by the electric spindle 24 begins to perform milling operations on the sample.

[0065] Optionally, the end mill 21 includes a shank and a cutting head. The shank portion of the end mill 21 is designed to mate with a second or first mounting cavity of a drive assembly (such as an electric spindle 24 or a pneumatic spindle 22). The diameter of the shank ranges from 2 to 6 mm, a size range that allows the end mill 21 to be adapted to a variety of different drive assemblies while maintaining sufficient strength and stability.

[0066] The cutting head of the milling cutter 21 directly participates in the milling operation. In this embodiment, the cutting head is made of diamond material, which has extremely high hardness and wear resistance, ensuring precise milling of the biological sample 5 under high-speed rotation.

[0067] The cutting tip material can be single-crystal diamond. Single-crystal diamond can be synthetic or natural. Synthetic single-crystal diamond is cost-effective and offers controllable crystal quality, while natural single-crystal diamond may have higher hardness and purity. A single-crystal diamond cutting tip ensures that the cutting edge remains sharp even at high speeds, thus providing a high-quality milled surface. The milling cutter 21 operates at speeds ranging from 30,000 to 80,000 rpm, ensuring the system can adapt to different milling requirements and biological sample types. Within this speed range, the milling cutter 21 provides sufficient milling force and surface finish while maintaining low vibration and noise levels.

[0068] In some embodiments, such as Figure 1 and Figure 3 As shown, the sample carrying module 1 is a three-dimensional precision platform, which includes an X-axis moving platform 11, a Y-axis moving platform 12, and a Z-axis moving platform 13 stacked sequentially from bottom to top. The biological sample 5 is placed on the Z-axis moving platform 13. The Z-axis moving platform 13 is used to drive the biological sample 5 to move along its height direction. The Y-axis moving platform 12 is used to drive the Z-axis moving platform 13 and the biological sample 5 to move along the width direction of the biological sample 5. The X-axis moving platform 11 is used to drive the Y-axis moving platform 12, the Z-axis moving platform 13, and the biological sample 5 to move along the length direction of the biological sample 5.

[0069] Specifically, the X-axis moving platform 11 includes a first base, a first guide rail, a first slider, and a first driving member. The first guide rail is fixed horizontally to the first base, the first slider is slidably connected to the first guide rail, and the first driving member is connected to the first slider. The first driving member provides power to drive the first slider to move along the first guide rail. The X-axis moving platform 11 is responsible for driving the Y-axis moving platform 12 (and the Z-axis moving platform 13 and biological sample 5 further above it) connected above it to move along the length direction of the biological sample 5. The Y-axis moving platform 12 is vertically disposed above the X-axis moving platform 11 and includes a second base, a second guide rail, a second slider, and a second driving member. The second base is fixed to the first slider, the second guide rail is fixed horizontally to the second base, the second slider is slidably connected to the second guide rail, and the second driving member is connected to the second slider. The second driving member provides power to drive the second slider to move along the second guide rail. The Y-axis moving platform 12 is responsible for driving the Z-axis moving platform 13 (and biological sample 5) connected above it to move along the width direction of the biological sample 5. Because it is fixed to the first slider of the X-axis moving platform 11, its movement is also controlled by the X-axis moving platform 11. The Z-axis moving platform 13 is vertically arranged above the Y-axis moving platform 12 and includes a third base, a third guide rail, a third slider, and a lifting drive assembly. The third base is fixed to the second slider, the third guide rail is fixed to the third base in the vertical direction, the third slider is slidably connected to the third guide rail, and the lifting drive assembly is connected to the third slider to provide power to drive the slider to move up and down along the guide rail. The Z-axis moving platform 13 is responsible for driving the biological sample 5 to move along its height direction. Because it is fixedly connected to both the Y-axis and X-axis moving platforms 11, its movement is jointly controlled by the X-axis and Y-axis moving platforms 12, achieving precise positioning in three-dimensional space.

[0070] Or, such as Figure 2 As shown, a Z-axis moving platform 13, an X-axis moving platform 11, and a Y-axis moving platform 12 are stacked sequentially from bottom to top. A biological sample 5 is mounted on the Y-axis moving platform 12. The Y-axis moving platform 12 drives the biological sample 5 to move along its width direction. The X-axis moving platform 11 drives the Y-axis moving platform 12 and the biological sample 5 to move along the length direction of the biological sample 5. The Z-axis moving platform 13 drives the X-axis moving platform 11, the Y-axis moving platform 12, and the biological sample 5 to move along the height direction of the biological sample 5. The structures of the Z-axis moving platform 13, the X-axis moving platform 11, and the Y-axis moving platform 12 are similar to those described above and can be referred to in the above embodiment; further details are omitted here.

[0071] Based on the above embodiments, in some embodiments, such as Figure 4As shown, the scanning module 3 is a TDI line-scan imaging device, which includes a TDI camera 31, a first illumination mechanism 32, a first detection mechanism 33, and a first objective lens. The first illumination mechanism 32 illuminates the section of the biological sample 5 through the first objective lens, and the TDI camera 31 and the first detection mechanism 33 capture the image of the section of the biological sample 5 through the first objective lens.

[0072] In this embodiment, the TDI camera 31 is capable of continuous exposure and integration along the scanning direction through its internal linear sensor array, which enhances the signal and reduces the noise. The first illumination mechanism 32 provides stable and uniform illumination, which is focused on the section of the biological sample 5 through the first objective lens. The first illumination mechanism 32 can employ a light source (e.g., LED, halogen lamp, or laser) and optical components (e.g., mirrors, lenses, etc.) to ensure that the light is uniformly illuminated on the sample. The first detection mechanism 33 is a sensor or detection element that works in conjunction with the TDI camera 31 to capture the image information of the section of the biological sample 5 imaged through the first objective lens.

[0073] When the scanning module 3 starts to work, the first illumination mechanism 32 provides uniform illumination to the section of the biological sample 5 through the first objective lens. The TDI camera 31 can be controlled to move along the pre-set scanning path (or the biological sample 5 itself moves), while its linear sensor array continuously exposes and integrates the section of the sample imaged through the first objective lens. The integrated image information is captured by the TDI camera 31 and converted into a digital signal for subsequent image processing and analysis.

[0074] Based on the above embodiment, in some embodiments, as shown, Figure 5 As shown, the scanning module 3 is a wide-field area imaging device, which includes an area camera 34, a second illumination mechanism 35, a second detection mechanism 36, and a second objective lens. The second illumination mechanism 35 illuminates the biological sample 5 through the second objective lens, and the area camera 34 and the second detection mechanism 36 capture the image of the section of the biological sample 5 through the second objective lens.

[0075] In this embodiment, the area camera 34 is equipped with a two-dimensional sensor array, which is capable of capturing the image information of the entire field of view at one time. The second illumination mechanism 35 provides uniform and sufficient intensity of illumination, which is uniformly illuminated on the biological sample 5 through the second objective lens. The second illumination mechanism 35 can employ a light source (e.g., LED array, halogen lamp array, or fiber-optic light source, etc.) and optical components (e.g., diffuser, mirrors, etc.) to ensure that the light can uniformly cover the entire sample area. The second detection mechanism 36 is a sensor array that works in conjunction with the area camera 34 to capture the image information of the section of the biological sample 5 imaged through the second objective lens.

[0076] When the scanning module 3 starts working, the second illumination mechanism 35 provides uniform illumination to the biological sample 5 through the second objective lens. The area array camera 34 captures a wide-angle image of the cross-section of the biological sample 5 through the second objective lens and converts it into a digital signal for storage and processing. The second detection mechanism 36 (i.e., the sensor array of the area array camera 34) is responsible for converting the captured light signal into an electrical signal, thereby generating a digital image.

[0077] Based on the above embodiments, in some embodiments, such as Figure 6 As shown, the scanning module 3 is a slant light imaging device, including: a third illumination mechanism 37, a third detection mechanism 38, a third objective lens and a fourth objective lens; the third illumination mechanism 37 illuminates one side of the cross section of the biological sample 5 at an angle through the third objective lens, and the third detection mechanism 38 takes pictures of the other side of the cross section of the biological sample 5 at an angle through the fourth objective lens.

[0078] In this embodiment, the third illumination mechanism 37 provides oblique illumination, directing light at a certain angle onto one side of the cross-section of the biological sample 5 through the third objective lens. The third illumination mechanism 37 may employ a light source (such as an LED, laser, etc.) and optical components (such as a mirror, prism, or lens, etc.) to ensure that the light can illuminate the sample at a predetermined angle and intensity. The third detection mechanism 38 is responsible for capturing image information of the other side of the cross-section of the biological sample 5 imaged by the fourth objective lens. The third detection mechanism 38 may include a camera (such as a CMOS or CCD camera) and related optical components (such as filters, lenses, etc.) to ensure that the obliquely imaged image can be clearly captured.

[0079] When the scanning module 3 starts working, the third illumination mechanism 37 illuminates one side of the cross-section of the biological sample 5 at a certain angle through the third objective lens. The light is scattered or reflected inside the sample, forming an oblique illumination area. The third detection mechanism 38 takes an image of the other side of the cross-section of the biological sample 5 through the fourth objective lens, capturing the image information after scattering or reflection inside the sample. The captured image information is converted into digital signals for storage and processing.

[0080] Based on the above embodiments, in some embodiments, such as Figure 7 As shown, the scanning module 3 is a vertical light sheet imaging device, including: a fourth illumination mechanism 30, a fifth illumination mechanism 39, a fourth detection mechanism 391, a fifth objective lens, a sixth objective lens, and a seventh objective lens; the fourth illumination mechanism 30 illuminates one side of the biological sample 5 vertically through the fifth objective lens, the fifth illumination mechanism 39 illuminates the other side of the biological sample 5 vertically through the sixth objective lens, and the fourth detection mechanism 391 captures the cross-section of the biological sample 5 vertically through the seventh objective lens.

[0081] In this embodiment, the fourth illumination mechanism 30 provides one side vertical illumination, and the light is vertically irradiated to one side of the biological sample 5 through the fifth objective lens. The fourth illumination mechanism 30 includes a light source (such as a laser light source) and related optical components (such as mirrors, lens groups, etc.) to ensure that the light can be irradiated to the sample in a vertical and uniform manner. The fifth illumination mechanism 39 provides the other side vertical illumination, and the light is vertically irradiated to the other side of the biological sample 5 through the sixth objective lens. The fifth illumination mechanism is similar to the fourth illumination mechanism 30. The fourth detection mechanism 391 is responsible for capturing the image information of the section of the biological sample 5 imaged through the seventh objective lens. The fourth detection mechanism 391 includes a high-sensitivity camera (such as a scientific-grade CMOS or CCD camera) and related optical components (such as filters, lens groups, etc.) to ensure that the image of the vertical light sheet imaging can be clearly captured.

[0082] When the scanning module 3 starts to work, the fourth illumination mechanism 30 and the fifth illumination mechanism 39 vertically irradiate light to both sides of the biological sample 5 through the fifth objective lens and the sixth objective lens, respectively. The light intersects inside the sample and forms a light sheet, illuminating a thin layer of the sample. The fourth detection mechanism 391 vertically photographs the section of the biological sample 5 illuminated by the light sheet through the seventh objective lens, and captures clear image information. The captured image information is converted into digital signals for storage and processing.

[0083] The utility model also provides a kind of control method of biological three-dimensional microscopic imaging system, and the structure of the biological three-dimensional microscopic imaging system can refer to Figures 1 to 7 Related written description.

[0084] As Figure 8 Indicated, the control method of the biological three-dimensional microscopic imaging system includes the following steps:

[0085] Step S810: making biological sample.

[0086] Step S820: set the biological sample in the sample carrying module, drive the biological sample to move to the milling station by the sample carrying module, and cooperate with the movement of the sample carrying module to mill the surface of the biological sample by the sample plane processing module.

[0087] Step S830: drive the biological sample to move to the milling station by the sample carrying module, drive the biological sample to raise a preset height by the sample carrying module, and cooperate with the movement of the sample carrying module to mill the biological sample by the sample plane processing module to form a smooth and mark-free surface on the section of the biological sample.

[0088] Step S840: drive the biological sample to move to the imaging station by the sample carrying module, and cooperate with the movement of the sample carrying module to image the smooth and mark-free surface of the biological sample by the scanning module to obtain the section image.

[0089] Step S850: alternately control the biological sample to be milled and imaged, so as to obtain a three-dimensional image of the biological sample according to a plurality of section images of the sequence.

[0090] In this embodiment, the biological sample 5 is first prepared. The biological sample is gradient dehydrated until completely dehydrated. The penetration of resin monomers and cross-linking agents is performed. The penetrated resin is polymerized at low / high temperature or ultraviolet to form a solid block. The resin material can be selected from natural resin or synthetic resin (such as epoxy resin, polyester resin, polyurethane resin, phenolic resin and acrylic resin), and can be prepared by free radical polymerization, ionic polymerization or polycondensation reaction to obtain the biological sample 5.

[0091] The prepared biological sample 5 is arranged on the sample carrying module 1. The sample carrying module 1 drives the biological sample 5 to move to the milling station, and the sample surface of the biological sample 5 is milled and flattened by the cooperation of the sample carrying module 1 and the sample surface processing module 2. For example, the sample carrying module 1 moves along the X and Y axes, and the sample surface processing module 2 mills, so as to remove the first layer of the biological sample 5 (usually about 0.1 mm thick) and realize the surface milling and flattening.

[0092] The sample carrying module 1 drives the biological sample 5 to move to the milling station and is lifted along the Z axis by a preset height (usually 1 micrometer). The sample carrying module 1 and the sample surface processing module 2 cooperate to mill the biological sample 5. For example, the sample carrying module 1 moves along the X and Y axes, and the sample surface processing module 2 mills to form a smooth and traceless section. The sample carrying module 1 drives the biological sample 5 to move to the imaging station. The scanning module 3 cooperates with the movement of the sample carrying module 1 to image the smooth and traceless surface of the biological sample 5. If the biological sample 5 is large, the sample carrying module 1 can be controlled to move along the X and Y axes, and the scanning module 3 can be controlled to take a picture to obtain a section image.

[0093] Then, the biological sample 5 is alternately controlled to be milled and imaged, that is, the plane of the biological sample 5 is milled again, and another section image at a preset height of the bottom of the section is obtained by imaging. Until a sufficient number of section images are obtained or the entire biological sample 5 is milled, the control processing module 4 reconstructs a three-dimensional image of the biological sample 5 according to a plurality of section images of the corresponding sequence.

[0094] The control method of the biological three-dimensional microscopic imaging system provided by the utility model, through the sample bearing module, the alternating movement of the biological sample between the milling station and the imaging station is accurately controlled, and the sample surface after milling is lifted, the continuity and accuracy of the imaging process are ensured. The milling cutter in the sample plane processing module can accurately mill the biological sample under the driving of the driving assembly, form a smooth surface without traces, and provide a high-quality cross section for subsequent scanning imaging. The scanning module performs high-resolution imaging on the smooth surface without traces, and the control processing module processes multiple cross-sectional images, which can reconstruct a high-precision three-dimensional image of the biological sample. The system has high automation, the cooperative work between the sample bearing module, the sample plane processing module and the scanning module can remove the surface of the biological sample at high speed, high smoothness, no trace and high precision, realize the rapid and continuous processing of the biological sample, and improve the imaging efficiency.

[0095] Finally, it should be noted that: the above examples are only used to illustrate the technical solutions of the utility model, and not to limit them; although the utility model has been described in detail with reference to the foregoing examples, those skilled in the art should understand that: it can still modify the technical solutions recorded in the foregoing examples, or make equivalent replacement for part of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the utility model.

Claims

1. A biological three-dimensional microscopic imaging system, characterized by, The application relates to a sample processing device, which comprises the following parts: a sample bearing module (1) suitable for placing a biological sample (5) and used for driving the biological sample (5) to move between a milling station and an imaging station alternately and driving the biological sample (5) to be lifted by a preset height after moving to the milling station; a sample planar processing module (2) arranged at the milling station and comprising a driving assembly and a milling cutter (21), wherein the driving end of the driving assembly is connected with the milling cutter (21), and the driving assembly is used for milling the biological sample (5) in cooperation with the movement of the sample bearing module (1) when the biological sample (5) moves to the milling station, so as to form a smooth surface on the section of the biological sample (5); a scanning module (3) arranged at the imaging station and used for imaging the smooth surface of the biological sample (5) in cooperation with the movement of the sample bearing module (1) when the biological sample (5) moves to the imaging station, so as to obtain a section image; a control processing module (4) electrically connected with the scanning module (3) and used for obtaining a plurality of section images according to the scanning module (3) and obtaining a three-dimensional image of the biological sample (5) according to the plurality of section images in sequence.

2. The biological three-dimensional microscopic imaging system of claim 1, wherein, The driving assembly comprises: an air compression motor; a pneumatic spindle (22) provided with a first installation cavity matched with the milling cutter (21) in the axial direction, wherein the pneumatic spindle (22) is driven to rotate at high speed by air pressure of the air compression motor; a first clamping piece (23) arranged at the opening end of the first installation cavity and provided with an adjustable first clamping structure, which is used for coaxially locking the milling cutter (21) with different diameter specifications on the rotation center axis of the pneumatic spindle (22).

3. The biological three-dimensional microscopic imaging system of claim 1, wherein, The driving assembly comprises: a driving motor; an electric spindle (24) connected with the output shaft of the driving motor through a shaft coupling, provided with a second installation cavity matched with the milling cutter (21) in the axial direction; a second clamping piece (25) arranged at the opening end of the second installation cavity and provided with an adjustable second clamping structure, which is used for coaxially locking the milling cutter (21) with different diameter specifications on the rotation center axis of the electric spindle (24).

4. The biological three-dimensional microscopic imaging system of claim 1, wherein, The milling cutter (21) comprises a cutter handle and a cutter head, wherein the diameter of the cutter handle is 2-6 mm, and the cutter head is a diamond cutter head; and the rotating speed of the milling cutter (21) is 30000-80000 revolutions per minute.

5. The bio-three-dimensional microscopic imaging system according to any one of claims 1-4, wherein, The sample bearing module (1) is a three-dimensional precision platform, and the three-dimensional precision platform comprises: The X-axis moving platform (11), the Y-axis moving platform (12) and the Z-axis moving platform (13) are sequentially stacked from bottom to top; the Z-axis moving platform (13) is provided with the biological sample (5), and the Z-axis moving platform (13) is used to drive the biological sample (5) to move along the height direction thereof; the Y-axis moving platform (12) is used to drive the Z-axis moving platform (13) and the biological sample (5) to move along the width direction of the biological sample (5); and the X-axis moving platform (11) is used to drive the Y-axis moving platform (12), the Z-axis moving platform (13) and the biological sample (5) to move along the length direction of the biological sample (5); Or, the Z-axis moving platform (13), the X-axis moving platform (11) and the Y-axis moving platform (12) are sequentially stacked from bottom to top; the Y-axis moving platform (12) is provided with the biological sample (5), and the Y-axis moving platform (12) is used to drive the biological sample (5) to move along the width direction thereof; the X-axis moving platform (11) is used to drive the Y-axis moving platform (12) and the biological sample (5) to move along the length direction of the biological sample (5); and the Z-axis moving platform (13) is used to drive the X-axis moving platform (11), the Y-axis moving platform (12) and the biological sample (5) to move along the height direction of the biological sample (5).

6. The biological three-dimensional microscopic imaging system according to any one of claims 1-4, wherein, The scanning module (3) is a TDI line scanning imaging device, comprising a TDI camera (31), a first illumination mechanism (32), a first detection mechanism (33) and a first objective lens; The first illumination mechanism (32) irradiates the section of the biological sample (5) through the first objective lens, and the TDI camera (31) and the first detection mechanism (33) shoot the section of the biological sample (5) through the first objective lens.

7. The bio-three-dimensional microscopic imaging system according to any one of claims 1-4, wherein, The scanning module (3) is a wide-field surface imaging device, comprising a surface array camera (34), a second illumination mechanism (35), a second detection mechanism (36) and a second objective lens; The second illumination mechanism (35) irradiates the biological sample (5) through the second objective lens, and the surface array camera (34) and the second detection mechanism (36) shoot the section of the biological sample (5) through the second objective lens.

8. The biological three-dimensional microscopic imaging system according to any one of claims 1-4, wherein, The scanning module (3) is an oblique light sheet imaging device, comprising a third illumination mechanism (37), a third detection mechanism (38), a third objective lens and a fourth objective lens; The third illumination mechanism (37) obliquely irradiates one side of the section on the biological sample (5) through the third objective lens, and the third detection mechanism (38) obliquely shoots the other side of the section on the biological sample (5) through the fourth objective lens.

9. The bio-three-dimensional microscopic imaging system according to any one of claims 1-4, wherein, The scanning module (3) is a vertical light sheet imaging device, comprising a fourth illumination mechanism (30), a fifth illumination mechanism (39), a fourth detection mechanism (391), a fifth objective lens, a sixth objective lens and a seventh objective lens; The fourth illumination mechanism (30) vertically irradiates one side of the biological sample (5) through the fifth objective lens, the fifth illumination mechanism (39) vertically irradiates the other side of the biological sample (5) through the sixth objective lens, and the fourth detection mechanism (391) vertically photographs the section of the biological sample (5) through the seventh objective lens.

Images