Stereoscopic spectral imaging microscope and method thereof

Abstract

The invention discloses a stereoscopic spectral imaging microscope and a method thereof, and belongs to the technical field of spectral imaging. The system is characterized by comprising a monochromatic source module, a microscope light path module and a spectral imaging module, the monochromatic source module comprises a light source, a monochromator, a monochromator scanning circuit and a monochromator motor; the microscope light path module comprises a collecting lens, an objective table, an objective table motor, an objective lens, a detector interface and a light shield. The spectral imaging module comprises a telescopic cylinder, a telescopic cylinder motor, a detector insertion cylinder, an area array photoelectric detector, a focusing circuit and an image plane scanning circuit. Compared with the prior art, the stereoscopic spectral imaging microscope and the method thereof provided by the invention have the advantages that the planar spectral imaging images from the first object plane to the Nth object plane of the sample to be detected can be sequentially obtained, and the stereoscopic spectral imaging image can be synthesized. And a researcher can research the shape, the defect, the physical structure and the chemical component of each part of the sample to be tested from a three-dimensional space.

Description

Technical Field

[0001] The present invention relates to the technical field of spectral imaging, and in particular to a stereo spectral imaging microscope and a method thereof. Background Art

[0002] A spectral imaging microscope is a microscope that can simultaneously obtain two-dimensional planar information and one-dimensional spectral information of a sample. The two-dimensional planar information can reveal the sample's shape, defects, and other qualities, while the one-dimensional spectral information can reveal the sample's internal physical structure and chemical composition. Therefore, spectral imaging microscopy has important applications in the study of microscopic structures such as cells. For example, spectral imaging microscopy can analyze the elemental composition of cells, identify different cells, distinguish between different cell growth stages, and measure the efficacy of drugs administered to cells. With the continuous development and improvement of spectral imaging microscopy technology, spectral imaging microscopy will play an even greater role in the biomedical field.

[0003] Any sample to be tested (including microscopic structures such as cells) has a finite volume and is three-dimensional. However, current spectral imaging microscopes can only produce two-dimensional spectral images of a specific physical plane of the sample, and cannot produce a three-dimensional spectral image of the sample. Because the shape, defects, physical structure, and chemical composition of different spatial components of the sample may vary, it is extremely necessary to develop a stereoscopic spectral imaging microscope that can produce three-dimensional spectral images. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a stereo spectral imaging microscope and method thereof, which enables researchers to study the shape, defects, physical structure and chemical composition of each spatial part of the sample to be tested in three-dimensional space.

[0005] The present invention adopts the following technical solutions to achieve the invention objectives: A stereo spectral imaging microscope and method thereof, characterized in that it comprises: a monochromatic light source module, a microscope optical path module and a spectral imaging module; the monochromatic light source module comprises a light source, a monochromator, a monochromator scanning circuit and a monochromator motor; the light source is installed above the light source entrance of the monochromator, the monochromator scanning circuit is installed in the monochromator, and the monochromator motor is installed on the monochromator cover of the monochromator; the microscope optical path module comprises a condenser, a stage, a stage motor, an objective lens, a detector interface and a light shield; the condenser, the stage and the objective lens are arranged in the light shield; the spectral imaging module comprises a telescopic cylinder, a telescopic cylinder motor, a detector plug The invention relates to a method for manufacturing a telescopic tube, a telescopic tube, an area array photoelectric detector, a focusing circuit and an image plane scanning circuit; the area array photoelectric detector is installed on the top of the inner cavity of the detector plug tube, the detector plug tube is fixedly connected to the telescopic tube, the detector plug tube and the telescopic tube are inserted into the detector interface, the telescopic tube motor is installed at the detector interface, and the focusing circuit and the image plane scanning circuit are installed in the inner cavity of the detector plug tube; the light emitted by the light source passes through the monochromator and is reflected by the condenser onto the sample to be tested on the stage, and the light passes through the sample to be tested and then passes through the objective lens, the detector interface, the telescopic tube, and the detector plug tube in sequence to form a real image on the area array photoelectric detector.

[0006] As a further limitation of the present technical solution, the light source includes a focusing concave mirror, a xenon lamp, a first convex lens, a second convex lens and a cooling fan. In order to reduce stray light, the xenon lamp is encapsulated in a light source cover coated with black.

[0007] As a further limitation of the present technical solution, the monochromator includes an entrance slit, a reflecting mirror, a concave mirror 1, a reflection grating, a concave mirror 2, an exit slit, a collimating lens, and a grating turntable. The entrance slit, the reflecting mirror, the concave mirror 1, the reflection grating, the concave mirror 2, the exit slit, the collimating lens, and the grating turntable are all encapsulated in a monochromator cover painted black.

[0008] As a further limitation of the present technical solution, the inner and outer diameters of the telescopic tube are equal to the inner and outer diameters of the detector insert tube, and the outer diameter of the telescopic tube and the outer diameter of the detector insert tube are equal to the inner diameter of the detector interface.

[0009] As a further limitation of the present technical solution, the monochromator scanning circuit includes a monochromator scanning control unit, a monochromator interface unit and the monochromator driving unit, and the monochromator interface unit, the monochromator scanning control unit, the monochromator driving unit and the monochromator motor are electrically connected in sequence.

[0010] As a further limitation of the present technical solution, the focusing circuit includes an image data processing unit, a focus value calculation unit, a focus algorithm unit, a stage control unit, a stage motor drive unit and an image data interface unit, and the array photodetector, the image data processing unit, the focus value calculation unit, the focus algorithm unit, the stage control unit, the stage motor drive unit and the stage motor are electrically connected in sequence.

[0011] As a further limitation of the present technical solution, the image plane scanning circuit includes a telescopic tube control unit, a telescopic tube motor drive unit and a telescopic tube interface unit, and the telescopic tube interface unit, the telescopic tube control unit, the telescopic tube motor drive unit and the telescopic tube motor are electrically connected in sequence.

[0012] A method for using a stereo spectral imaging microscope, characterized by comprising the following steps: S1: Preparation before measurement; The process includes turning on the device, the monochromatic light source module enters a self-test state and a preheating state; placing the sample to be tested on the stage and closing the light shield; adjusting the stage so that the bottom object distance of the sample to be tested is less than 1 times the focal length of the objective lens; adjusting the extension length of the telescopic tube to zero; setting the wavelength scanning range. l 1~ l 2. Wavelength resolution Δ l and image plane scan number N ; S2: imaging focus; Click to start focusing, the light source first outputs a wavelength of l 1, the focusing circuit starts searching, the stage motor drives the stage to change its height to adjust the object distance for focusing, and as the stage height changes, the object distance gradually increases, and the real image of the first object plane of the sample to be measured gradually approaches the area array photoelectric detector from a distance and is searched by the area array photoelectric detector. Focusing is completed when it reaches the clearest point; S3: Obtain spectral imaging images of each plane; After the image plane scanning circuit 37 receives the image plane scanning instruction from the computer, the array photoelectric detector 33 moves to the first image plane, the second image plane, the third image plane, ..., until the third image plane. N At the image plane, the plane spectrum imaging image of the first object plane, the plane spectrum imaging image of the second object plane, the plane spectrum imaging image of the third object plane, ..., are obtained in sequence until the plane spectrum imaging image of the first object plane, the plane spectrum imaging image of the second object plane, the plane spectrum imaging image of the third object plane, ..., are obtained. N Planar spectral imaging image of the object plane; S4: Synthetic stereo spectral imaging image; The stereo spectral imaging image is composed of the aboveN The plane spectral imaging images are sequentially stacked and fitted to synthesize. Since the object distances of the object planes are different, the magnifications of the plane spectral imaging images of the object planes are different. After the computer performs appropriate scaling correction on the plane spectral imaging images of the object planes, they are sequentially stacked and fitted to synthesize the stereo spectral imaging image of the sample to be tested.

[0013] Compared with the prior art, the advantages and positive effects of the present invention are: 1. When the area array photoelectric detector is in a certain image plane, it can obtain a planar spectral imaging image of the sample to be tested, realizing the function of the current spectral imaging microscope.

[0014] 2. When the array photoelectric detector moves from the first image plane of the sample to be tested to the N When the image plane is formed, the image data from the first object plane to the first object plane of the sample to be tested can be obtained in sequence. N The two-dimensional spectral imaging image of the object plane is then sequentially superimposed and fitted to synthesize the three-dimensional spectral imaging image of the sample to be tested. This allows researchers to study the shape, defects, physical structure, and chemical composition of each part of the sample to be tested in three-dimensional space. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 It is a structural schematic diagram of the present invention.

[0016] Figure 2 This is a diagram showing the locations of the main components of the present invention.

[0017] Figure 3 This is a logic structure diagram of the monochromator scanning circuit of the present invention.

[0018] Figure 4 This is a logic structure diagram of the focusing circuit of the present invention.

[0019] Figure 5 This is a logic structure diagram of the image plane scanning circuit of the present invention.

[0020] Figure 6 This is a schematic diagram of the stereo spectral imaging principle of the present invention.

[0021] Figure 7 Schematic diagram of the spectral imaging method of the present invention.

[0022] In the figure: 1. Monochromatic light source module; 11. Light source; 11a. Concave mirror; 11b. Xenon lamp; 11c. Convex lens 1; 11d. Convex lens 2; 11e. Light source cover; 12. Monochromator; 12a. Entrance slit; 12b. Reflector; 12c. Concave mirror 1; 12d. Reflection grating; 12e. Concave mirror 2; 12f. Exit slit; 12g. Collimating lens; 12h. Monochromator cover; 12i. Grating turntable; 13. Monochromator scanning circuit; 13a. Monochromator scanning control unit; 13b. Monochromator interface unit; 13c. Monochromator drive unit; 14. Monochromator motor; 2. Microscope optical path module; 21. Condenser; 22. Stage; 23. Sample to be tested 1. Product; 24. Objective lens; 25. Detector interface; 26. Light shield; 27. Stage motor; 3. Spectral imaging module; 31. Telescopic tube; 31a. Rack; 32. Detector insert; 33. Array photodetector; 34. Real image of the sample to be measured; 35. Telescopic tube motor; 35a. Gear; 36. Focusing circuit, 36a. Image data processing unit, 36b. Focus value calculation unit, 36c. Focusing algorithm unit, 36d. Stage control unit, 36e. Stage motor drive unit, 36f. Image data interface unit; 37. Image plane scanning circuit, 37a. Telescopic tube control unit, 37b. Telescopic tube motor drive unit, 37c. Telescopic tube interface unit. DETAILED DESCRIPTION

[0023] A specific embodiment of the present invention is described in detail below with reference to the accompanying drawings, but it should be understood that the protection scope of the present invention is not limited by the specific embodiment.

[0024] The present invention comprises: a monochromatic light source module 1, a microscope optical path module 2 and a spectral imaging module 3; the monochromatic light source module 1 comprises a light source 11, a monochromator 12, a monochromator scanning circuit 13 and a monochromator motor 14; the light source 11 is installed above the light source entrance of the monochromator 12, the monochromator scanning circuit 13 is installed in the monochromator 12, and the monochromator motor 14 is installed on the monochromator cover 12h of the monochromator 12; the microscope optical path module 2 comprises a condenser 21, a stage 22, a stage motor 27, an objective lens 24, a detector interface 25 and a light shield 26; the entire microscope base and the outside of the fuselage are installed with a black inner wall. , the light shield 26 can be opened and closed, the condenser 21, the stage 22 and the objective lens 24 are arranged in the light shield 26; the height of the stage 22 can be changed; the stage motor 27 is installed in the original fine-focusing knob position in the light shield 26, replacing the original fine-focusing knob, and is connected to the stage 22 by connecting the lifting screw and other existing technologies, so that when the stage motor 27 rotates, the height of the stage 22 can be changed; the monochromator 12 is fixedly connected to the light shield 26; the spectral imaging module 3 includes a telescopic cylinder 31, a telescopic cylinder motor 35, a detector plug 32, and an array photoelectric detector 33 , focusing circuit 36 ​​and image plane scanning circuit 37; the area array photoelectric detector 33 is a monochrome CCD image sensor (or monochrome CMOS image sensor), which is installed on the top of the inner cavity of the detector plug tube 32, and the detector plug tube 32 is fixedly connected to the telescopic tube 31. The detector plug tube 32 and the telescopic tube 31 are inserted into the detector interface 25, and the detector plug tube 32 and the telescopic tube 31 can move along the axial direction of the detector interface 25. The telescopic tube motor 35 is installed at the detector interface 25, and the rotor output shaft of the telescopic tube motor 35 is installed with a gear 35a, and a rack is installed on one side of the telescopic tube 31. 31a, the gear 35a and the rack 31a are engaged with each other, so that when the telescopic cylinder motor 35 rotates, the gear 35a pushes the rack 31a to move up and down, driving the telescopic cylinder 31 to move up and down; the inner cavity of the detector insert 32 is installed with the focusing circuit 36 ​​and the image plane scanning circuit 37; the light emitted by the light source 11 passes through the monochromator 12 and is reflected by the condenser 21 onto the sample to be measured 23 on the stage 22. After passing through the sample to be measured 23, the light passes through the objective lens 24, the detector interface 25, the telescopic cylinder 31, and the detector insert 32 in sequence to form a real image on the area array photodetector 33.

[0025] There is no filter in front of the photosensitive unit of the monochrome CCD image sensor (or monochrome CMOS image sensor), which can pass monochromatic light of any wavelength, and thus can capture monochrome images of any wavelength. These image data are the monochrome layers in the plane spectral imaging image. When the monochromatic light source is within the set wavelength scanning range, l 1~ l 2 according to the set wavelength resolution Δ l During scanning, each time the wavelength changes, the area array photodetector obtains a set of image data at that wavelength. After the scan is complete, hundreds of sets of image data at different wavelengths can be obtained, and these hundreds of images at different wavelengths can be synthesized into a single planar spectral imaging image.

[0026] The light source 11 includes a focusing concave mirror 11a, a xenon lamp 11b, a convex lens 11c, a convex lens 2 11d and a cooling fan. In order to reduce stray light, the xenon lamp 11b is enclosed in a light source cover 11e coated with black.

[0027] The monochromator 12 is a Czerny-Turner monochromator, including an entrance slit 12a, a reflector 12b, a concave mirror 12c, a reflection grating 12d, a concave mirror 12e, an exit slit 12f, a collimating lens 12g, and a grating turntable 12i. To reduce stray light, the entrance slit 12a, the reflector 12b, the concave mirror 12c, the reflection grating 12d, the concave mirror 12e, the exit slit 12f, the collimating lens 12g, and the grating turntable 12i are all enclosed in a black-coated monochromator cover 12h.

[0028] The inner and outer diameters of the telescopic tube 31 are equal to the inner and outer diameters of the detector insert tube 32 , and the outer diameters of the telescopic tube 31 and the detector insert tube 32 are equal to the inner diameter of the detector interface 25 . In order to reduce stray light, the inner wall of the telescopic tube 31 , the inner wall of the detector insert tube 32 , and the detector interface 25 are painted black.

[0029] The monochromator scanning circuit 13 includes a monochromator scanning control unit 13a, a monochromator interface unit 13b, and a monochromator drive unit 13c. The monochromator interface unit 13b, the monochromator scanning control unit 13a, the monochromator drive unit 13c, and the monochromator motor 14 are electrically connected in sequence. After receiving computer instructions, the monochromator scanning control unit 13a outputs a control signal to the monochromator drive unit 13c. The monochromator drive unit 13c converts the digital signal output by the monochromator scanning control unit 13a into an appropriate voltage / current power signal to drive the monochromator motor 14 to rotate in a specific direction. When the monochromator motor 14 rotates, the reflection grating 12d undergoes angular deflection, resulting in monochromatic light of different wavelengths at the exit slit 12f. The monochromator interface unit 13b is responsible for transmitting wavelength data to the computer and receiving self-test or scanning instructions from the computer.

[0030] In the Czerny-Turner monochromator, the reflection grating 12d is mounted on a grating turntable 12i inside a monochromator cover 12h, and the output shaft of the monochromator motor 14 mounted on the monochromator cover 12h is connected to the grating turntable 12i.

[0031] The focusing circuit 36 ​​includes an image data processing unit 36a, a focus value calculation unit 36b, a focus algorithm unit 36c, a stage control unit 36d, a stage motor drive unit 36e and an image data interface unit 36f. The array photodetector 33, the image data processing unit 36a, the focus value calculation unit 36b, the focus algorithm unit 36c, the stage control unit 36d, the stage motor drive unit 36e and the stage motor 27 are electrically connected in sequence. After receiving a focus command from the computer, the focusing circuit 36 ​​begins searching. The area array photodetector 33 converts the collected light signal into an electrical signal and transmits it to the image data processing unit 36a. The signal is then processed by the focus value calculation unit 36b and the focus algorithm unit 36c. Based on the focus value calculation result, the stage control unit 36d outputs a control signal to the stage motor drive unit 36e. The stage motor drive unit 36e converts the digital signal output by the stage control unit 36d into an appropriate voltage / current power signal to drive the stage motor 27. The stage motor 27 drives the height of the stage 22 to adjust the object distance for focusing. The image data interface unit 36f (one end electrically connected to the image data processing unit 36a and the other end electrically connected to the computer) is responsible for transmitting the image data processed by the image data processing unit 36a and stage 22 height information to the computer and receiving focus commands from the computer.

[0032] The image plane scanning circuit 37 includes a telescopic tube control unit 37a, a telescopic tube motor drive unit 37b, and a telescopic tube interface unit 37c. The telescopic tube interface unit 37c, the telescopic tube control unit 37a, the telescopic tube motor drive unit 37b, and the telescopic tube motor 35 are electrically connected in sequence. After the image plane scanning circuit 37 receives an image plane scanning instruction from the computer, the telescopic tube control unit 37a outputs a control signal to the telescopic tube motor drive unit 37b. The telescopic tube motor drive unit 37b converts the digital signal output by the telescopic tube control unit 37a into an appropriate voltage / current power signal to drive the telescopic tube motor 35. The telescopic tube motor 35 drives the telescopic tube 31 to the specified image plane. The telescopic tube interface unit 37c is responsible for transmitting the position information of the telescopic tube 31 to the computer and receiving the image plane scanning instruction from the computer.

[0033] The principle of stereo spectral imaging implemented by the present invention can be found in Figure 6 , Figure 6 The following is a diagram showing the principle of stereo spectral imaging of a stereo spectral imaging microscope according to an embodiment of the present invention. Figure 6 The three-dimensional letter "H" is used to draw the sample 23 to be tested. The sample 23 to be tested has a certain thickness along the main optical axis. The sample 23 to be tested with a certain thickness has an infinite number of object planes along the main optical axis. Figure 6 Only five object planes are drawn, corresponding to five image planes. The real image of an object point on the first object plane after the objective lens 24 is located on the first image plane; the real image of an object point on the second object plane after the objective lens 24 is located on the second image plane; ...; the real image of an object point on the fifth object plane after the objective lens 24 is located on the fifth image plane. When the array photoelectric detector 33 moves to the first object plane, the real image of the object point on the fifth object plane is located on the fifth image plane. i When the image plane is on, start the monochromatic light source 1 to scan the wavelength within the set range. l 1~ l 2 according to the set wavelength resolution Δ l Start scanning. Every time the monochromatic light source 1 changes its wavelength, the array photodetector 33 obtains a set of image data of the wavelength. After the scanning is completed, a total of [( l 2- l 1) / Δ l ] sets of image data with different wavelengths, and the image data with different wavelengths can be synthesized into the first i The image plane corresponds to i If the above method is used to sequentially obtain the plane spectral imaging image of the first object plane, the plane spectral imaging image of the second object plane, ..., until the plane spectral imaging image of the first object plane, the plane spectral imaging image of the second object plane, ..., until the plane spectral imaging image of the second object plane, ... NThe plane spectral imaging image of the object plane can be used to synthesize a stereoscopic spectral imaging image. Synthesizing a stereoscopic spectral imaging image does not require an infinite number of plane spectral imaging images of the sample 23 to be tested. Since the thickness of the sample 23 to be tested along the principal optical axis is very small, generally, the plane spectral imaging images of five object planes stacked and fitted sequentially can meet the accuracy requirements. Because the object distance and image distance of the plane spectral imaging images of each object plane are different, the magnification of the plane spectral imaging images of each object plane is different. After the computer scales and corrects the plane spectral imaging images of each object plane, they are stacked and fitted sequentially to synthesize a stereoscopic spectral imaging image of the sample 23 to be tested.

[0034] A method for using a stereo spectral imaging microscope comprises the following steps: S1: Preparation before measurement; The process includes turning on the power, the monochromatic light source module 1 enters the self-test state and the preheating state; placing the sample 23 to be tested on the stage 2, closing the light shield 26; adjusting the stage 22 so that the bottom object distance of the sample 23 to be tested is less than 1 times the focal length of the objective lens 24; adjusting the extension length of the telescopic tube 31 to zero; setting the wavelength scanning range. l 1~ l 2. Wavelength resolution Δ l and image plane scan number N ; The specific implementation process is as follows: S11: Turn on the power, and the monochromatic light source module 1 enters a self-test state. The monochromatic light source module 1 automatically calibrates the wavelength and performs self-tests on the system parameters and dark current. After the self-test is completed, the monochromatic light source module 1 enters a preheating state. The preheating time is 20 minutes. After the preheating is completed, the monochromatic light source module 1 automatically detects the dark current once again. S12: Open the light shield 26 of the microscope, place the sample 23 to be measured on the stage 22, and close the light shield 26; adjust the stage 22 so that the bottom object distance of the sample 23 to be measured is less than 1 times the focal length of the objective lens 24; adjust the extension length of the telescopic tube 31 to zero; S13: The wavelength scanning range set in this embodiment is 400nm~1000nm; the wavelength resolution set in this embodiment is 8nm; S14: In this embodiment, the number of image plane scans is set to 5, and the bottom layer of the sample 23 to be tested is used as the first object plane of the sample 23 to be tested, and the top layer of the sample 23 to be tested is used as the fifth object plane of the sample 23 to be tested; S2: imaging focus; Click to start focusing, the light source 11 first outputs a wavelength of l1, the focusing circuit 36 ​​starts searching, the area array photoelectric detector 33 converts the collected light signal into an electrical signal and transmits it to the image data processing unit 36a, which is processed by the focus value calculation unit 36b and the focus algorithm unit 36c in sequence. According to the focus value calculation result, the stage control unit 36d outputs a control signal to the stage motor driving unit 36e, and the stage motor driving unit 36e converts the digital signal output by the stage control unit 36d into a suitable voltage / current power signal to drive the stage motor 27. The stage motor 27 drives the height of the stage 22 to adjust the object distance for focusing. As the height of the stage 22 changes, the object distance gradually increases, and the real image of the first object plane of the sample to be measured 23 gradually approaches the area array photoelectric detector 33 from a distance and is searched by the area array photoelectric detector 33. The focusing is completed when it reaches the clearest point. The specific automatic focusing implementation process is as follows: S21: The focusing circuit 36 ​​starts searching, the stage motor 27 drives the stage 22 to change its height, and the object distance of the sample 23 to be measured gradually increases. As the object distance exceeds 1 times the focal length, the real image of the first object plane of the sample 23 to be measured gradually approaches the area array photodetector 33 from a distance and is detected by the area array photodetector 33. S22: The area array photoelectric detector 33 outputs the original image data of the real image of the first object plane of the sample to be tested 23 to the image data processing unit 36a. The image data processing unit 36a divides the obtained original image data into n sub-region image data according to a preset image data processing algorithm, and sends the obtained n image data to the focus value calculation unit 36b. S23: The focus value calculation unit 36b calculates focus values ​​for the image data of the n sub-regions respectively, and obtains n different sets of high-frequency component focus values ​​and low-frequency component focus values; S24: The focusing algorithm unit 36c selects an appropriate value as a focus evaluation value based on the n sets of focus values ​​and the threshold value. The focus evaluation value is used to determine the region where the optimal focus value is located. S25: Based on the selected focus evaluation value, a coarse focus search algorithm with an adaptive step size is used for searching. During the coarse focus search algorithm, the stage control unit 36d sends a drive signal to the stage motor 27, so that the stage motor 27 can control the stage 22 to complete m round trips to find the area with the best focus value. If m round trips are completed, the process proceeds to the next step S26; otherwise, the process returns to the previous step S24. S26: Reading a certain position A0 within the range of the optimal focus value determined when running the coarse focus search algorithm, the stage control unit 36d sends a drive signal to the stage motor 27. After receiving the drive signal, the stage motor 27 controls the stage 22 to move to the aforementioned position A0; S27: running a fine focus search algorithm using a fixed step size according to the focus evaluation value selected in step S24; S28: During the fine focus search process, the stage control unit 36d sends a drive signal to the stage motor 27. After receiving the drive signal, the stage motor 27 controls the stage 22 to complete k round trips to find the position A1 of the stage 22 when the optimal focus value is obtained. If k round trips are completed, the next step S29 is executed; otherwise, the process returns to the previous step S27. S29: Reading the position A1 of the stage 22 when the best focus value is obtained during the fine focus search process, the stage control unit 36d sends a drive signal to the stage motor 27. After receiving the drive signal, the stage motor 27 controls the stage 22 to move to the position A1 of the stage 22 when the best focus value is obtained. S210: The fine focus search process is completed. At this point, the focus adjustment is completed, and the real image of the first object plane of the sample 23 to be measured stops at the position of the area array photoelectric detector 33; S3: Obtain spectral imaging images of each plane; After the image plane scanning circuit 37 receives the image plane scanning instruction from the computer, the array photoelectric detector 33 moves to the first image plane, the second image plane, the third image plane, ..., until the third image plane. N At the image plane, the plane spectrum imaging image of the first object plane, the plane spectrum imaging image of the second object plane, the plane spectrum imaging image of the third object plane, ..., are obtained in sequence until the plane spectrum imaging image of the first object plane, the plane spectrum imaging image of the second object plane, the plane spectrum imaging image of the third object plane, ..., are obtained. N Planar spectral imaging image of the object plane; The automatic process of obtaining the spectral imaging images of each plane of the sample 23 to be tested is as follows: S31: After focusing is completed, the area array photodetector 33 stops at the clear real image position of the first object plane of the sample 23 to be tested. Each photosensitive unit on the area array photodetector 33 converts the light signal at that point into an electrical signal. These electrical signals are processed by the image data processing unit 36a and then transmitted to the computer by the image data interface unit 36f. The computer can then obtain 400nm image data of the first object plane of the sample 23 to be tested. S32: After the computer obtains the 400nm image data of the first object plane of the sample 23 to be tested, the computer sends a command to the monochromator 12. The monochromator 12 outputs monochromatic light with a wavelength of 408nm. At this time, each photosensitive unit on the area array photodetector 33 converts the light signal at that point into an electrical signal. These electrical signals are processed by the image data processing unit 36a and then transmitted to the computer by the image data interface unit 36f. The computer thus obtains the 408nm image data of the first object plane of the sample 23 to be tested. S33: After the computer obtains the 408nm image data of the first object plane of the sample 23 to be tested, the computer sends a command to the monochromator 12. The monochromator 12 outputs monochromatic light with a wavelength of 416nm. At this time, each photosensitive unit on the area array photodetector 33 converts the light signal at that point into an electrical signal. These electrical signals are processed by the image data processing unit 36a and then transmitted to the computer by the image data interface unit 36f. The computer thus obtains the 416nm image data of the first object plane of the sample 23 to be tested. S34: Similarly, the computer can also obtain 424 nm image data, 430 nm image data, ..., up to 1000 nm image data of the first object plane of the sample 23 to be tested, for a total of 75 sets; S35: The computer synthesizes the 75 sets of image data at different wavelengths to obtain a planar spectral imaging image of the first object plane of the sample 23 to be tested; S36: When the spectral imaging scan of the first object plane of the sample 23 is completed, the computer sends a spectral imaging scan instruction for the second object plane of the sample 23 to the image plane scanning circuit 37. The image plane scanning circuit 37 drives the extension of the telescopic cylinder 31 to gradually increase, and the area array photodetector 33 moves to the real image position of the second object plane of the sample 23. According to the above methods of S31-S35, a planar spectral imaging image of the second object plane of the sample 23 is obtained. S37: Similarly, according to the above S36 method, the third object plane, the fourth object plane, ..., the fourth object plane of the sample 23 to be tested are obtained. N -1 object plane, N Planar spectral imaging image of the object plane; S4: Synthetic stereo spectral imaging image; The stereo spectral imaging image is composed of the above N The plane spectral imaging images are sequentially stacked and fitted to synthesize. Since the object distances of the object planes are different, the magnifications of the plane spectral imaging images of the object planes are different. After the computer performs appropriate scaling correction on the plane spectral imaging images of the object planes, they are sequentially stacked and fitted to synthesize the stereo spectral imaging image of the sample to be tested 23.

[0035] If the wavelength scanning range is l 1~ l2. Wavelength resolution is Δ l , the number of image plane scans is N , then the stereo spectral imaging image is composed of N The plane spectral imaging images are sequentially stacked and fitted. Based on the stereo spectral imaging images, researchers can study the shape and defects of each layer of the sample 23 in three-dimensional space. When the computer mouse moves to a layer of the stereo spectral imaging image, the computer can display the plane spectral imaging image corresponding to the layer; since any plane spectral imaging image is composed of [( l 2- l 1) / Δ l ] Group from l 1 to l 2 is processed using image data at different wavelengths. When the computer mouse moves to a point on the planar spectral imaging image, the computer can obtain the corresponding spectral curve. Based on the spectral curve, researchers can study the physical structure and chemical composition of the sample under test.

[0036] Because the wavelength scanning range set in this embodiment is 400nm to 1000nm, the wavelength resolution is 8nm, and the number of image plane scans is 5, the stereoscopic spectral imaging image obtained in this embodiment is a composite of five planar spectral imaging images stacked and fitted in sequence. Each planar spectral imaging image is obtained by processing 75 sets of image data at different wavelengths ranging from 400nm to 1000nm.

[0037] The above disclosure is only a specific embodiment of the present invention, but the present invention is not limited thereto. Any changes that can be conceived by those skilled in the art should fall within the scope of protection of the present invention.

Claims

1. A stereo spectral imaging microscope, characterized in that: include: A monochromatic light source module (1), a microscope optical path module (2) and a spectral imaging module (3); The monochromatic light source module (1) comprises a light source (11), a monochromator (12), a monochromator scanning circuit (13) and a monochromator motor (14); The light source (11) is installed above the light source entrance of the monochromator (12), the monochromator scanning circuit (13) is installed in the monochromator (12), and the monochromator motor (14) is installed on the monochromator cover (12h) of the monochromator (12); The microscope optical path module (2) comprises a condenser (21), a stage (22), a stage motor (27), an objective lens (24), a detector interface (25) and a light shield (26); The condenser (21), the stage (22) and the objective lens (24) are arranged in the light shield (26); The spectral imaging module (3) comprises a telescopic tube (31), a telescopic tube motor (35), a detector insert tube (32), a planar array photoelectric detector (33), a focusing circuit (36) and an image plane scanning circuit (37); The planar array photoelectric detector (33) is mounted on the top of the inner cavity of the detector insert tube (32); the detector insert tube (32) is fixedly connected to the telescopic tube (31); the detector insert tube (32) and the telescopic tube (31) are inserted into the detector interface (25); the telescopic tube motor (35) is mounted at the detector interface (25); and the focusing circuit (36) and the image plane scanning circuit (37) are mounted in the inner cavity of the detector insert tube (32); The light emitted by the light source (11) passes through the monochromator (12) and is reflected by the condenser (21) onto the sample to be tested (23) on the stage (22). The light passes through the sample to be tested (23) and then sequentially passes through the objective lens (24), the detector interface (25), the telescopic tube (31), and the detector insert tube (32) to form a real image on the array photoelectric detector (33).

2. The stereo spectral imaging microscope according to claim 1, characterized in that: The light source (11) comprises a focusing concave mirror (11a), a xenon lamp (11b), a convex lens 1 (11c), a convex lens 2 (11d) and a heat dissipation fan. In order to reduce stray light, the xenon lamp (11b) is encapsulated in a light source cover (11e) coated in black.

3. The stereo spectral imaging microscope according to claim 1, characterized in that: The monochromator (12) comprises an incident slit (12a), a reflector (12b), a concave mirror 1 (12c), a reflection grating (12d), a concave mirror 2 (12e), an exit slit (12f), a collimating lens (12g), and a grating turntable (12i); the incident slit (12a), the reflector (12b), the concave mirror 1 (12c), the reflection grating (12d), the concave mirror 2 (12e), the exit slit (12f), the collimating lens (12g), and the grating turntable (12i) are all encapsulated in a monochromator cover (12h) coated in black.

4. The stereo spectral imaging microscope according to claim 1, characterized in that: The inner and outer diameters of the telescopic tube (31) are equal to the inner and outer diameters of the detector insert tube (32); the outer diameter of the telescopic tube (31) and the outer diameter of the detector insert tube (32) are equal to the inner diameter of the detector interface (25).

5. The stereo spectral imaging microscope according to claim 1, characterized in that: The monochromator scanning circuit (13) comprises a monochromator scanning control unit (13a), a monochromator interface unit (13b) and a monochromator driving unit (13c); the monochromator interface unit (13b), the monochromator scanning control unit (13a), the monochromator driving unit (13c) and the monochromator motor (14) are electrically connected in sequence.

6. The stereo spectral imaging microscope according to claim 1, characterized in that: The focusing circuit (36) comprises an image data processing unit (36a), a focus value calculation unit (36b), a focus algorithm unit (36c), a stage control unit (36d), a stage motor drive unit (36e) and an image data interface unit (36f); the array photodetector (33), the image data processing unit (36a), the focus value calculation unit (36b), the focus algorithm unit (36c), the stage control unit (36d), the stage motor drive unit (36e) and the stage motor (27) are electrically connected in sequence.

7. The stereo spectral imaging microscope according to claim 1, characterized in that: The image plane scanning circuit (37) comprises a telescopic tube control unit (37a), a telescopic tube motor driving unit (37b) and a telescopic tube interface unit (37c); the telescopic tube interface unit (37c), the telescopic tube control unit (37a), the telescopic tube motor driving unit (37b), and the telescopic tube motor (35) are electrically connected in sequence.

8. The method for using the stereo spectral imaging microscope according to claim 1, characterized in that: The following steps are involved: S1: Preparation before measurement; The method comprises the steps of turning on the machine, wherein the monochromatic light source module (1) enters a self-test state and a preheating state; placing a sample to be tested (23) on the stage (2) and closing the light shield (26); adjusting the stage (22) so that the bottom object distance of the sample to be tested (23) is less than 1 times the focal length of the objective lens (24); adjusting the extension length of the telescopic tube (31) to zero; and setting the wavelength scanning range. λ 1~ λ 2. Wavelength resolution Δ λ and image plane scan number N ; S2: imaging focus; Click to start focusing, the light source (11) first outputs a wavelength of λ 1, the focusing circuit (36) starts searching, the stage motor (27) drives the stage (22) to change its height to adjust the object distance for focusing, and as the height of the stage (22) changes, the object distance gradually increases, and the real image of the first object plane of the sample to be tested (23) gradually approaches the area array photoelectric detector (33) from a distance and is searched by the area array photoelectric detector (33), and the focusing is completed when the image is clearest; S3: Obtain spectral imaging images of each plane; After the image plane scanning circuit (37) receives the image plane scanning instruction sent by the computer, the array photoelectric detector (33) moves to the first image plane, the second image plane, the third image plane, ..., until the third image plane. N At the image plane, a plane spectral imaging image of the first object plane, a plane spectral imaging image of the second object plane, a plane spectral imaging image of the third object plane, ..., is obtained in sequence until a plane spectral imaging image of the first object plane, a plane spectral imaging image of the second object plane, a plane spectral imaging image of the third object plane, ..., is obtained. N Planar spectral imaging image of the object plane; S4: Synthetic stereo spectral imaging image: The stereo spectral imaging image is composed of the above N The plane spectral imaging images are sequentially stacked and fitted to synthesize. Since the object distances of the object planes are different, the magnifications of the plane spectral imaging images of the object planes are different. After the computer performs appropriate scaling correction on the plane spectral imaging images of the object planes, they are sequentially stacked and fitted to synthesize a three-dimensional spectral imaging image of the sample to be tested (23).

Images