A label-free photoacoustic pathology microscopy imaging system and imaging method

Abstract

The present invention discloses a label-free photoacoustic pathology microscopic imaging system and imaging method. The system includes: an optical surface wave sensor for carrying tissue samples and water layers; an excitation light generating device; a detection light generating device; an objective lens, which allows the excitation light beam and the detection light beam to be incident on the tissue sample through the optical surface wave sensor; a processing and control device, which is used to control the movement of the excitation light beam to scan the tissue sample; an optical surface wave sensor, which is used to fully reflect the detection light beam, so that the reflected detection light beam is emitted through the objective lens; a processing and control device, which is used to split the detection light beam, synthesize photoacoustic signals and reconstruct images to obtain a target image. The imaging system has a high degree of integration and integration, and uses the optical surface wave sensor as a slide to carry the tissue sample, which can retain high-frequency signals to a great extent and improve the longitudinal resolution. In addition, the excitation light beam and the detection light beam share an objective lens, which can better ensure the coaxiality of the two light beams.

Description

Technical Field

[0001] The present invention relates to the field of microscopic imaging, and in particular to a label-free photoacoustic pathology microscopic imaging system and an imaging method. Background Art

[0002] At present, photoacoustic microscopy technology is the fastest-growing emerging biological imaging method in the past decade, and it has great potential in pathological detection. Liu Wenzhong's team at Northwestern University in the United States used photoacoustic imaging technology to detect the structure of mouse eyes. Compared with other technologies, it can better detect retinal blood oxygen saturation and melanin concentration, further promoting the clinical transfer of photoacoustic imaging. Catherine Martel's team in the United States combined photoacoustic microscopy with microelectromechanical system scanning mirrors for lymphatic pathology detection. Although photoacoustic microscopy technology has made great progress, it is restricted by the properties of piezoelectric materials. Photoacoustic systems generally have the defects of narrow detection bandwidth and low sensitivity. The narrow detection bandwidth leads to low longitudinal resolution of photoacoustic imaging, which in turn causes inaccurate positioning in the depth direction, which is not conducive to actual pathological imaging. In order to solve this problem, optical surface wave-based sensing technology is applied to the field of photoacoustic imaging. By detecting the phase change of the detection light, the high sensitivity of the photoacoustic imaging system is achieved.

[0003] Compared with traditional photoacoustic sensors, optical surface wave sensors have the advantages of high sensitivity and broadband photoacoustic detection. In the emerging field of photoacoustic imaging based on optical surface wave sensing, the University of Virginia in the United States has established a photoacoustic microscopy system using plasma detection technology, which has achieved high spatial resolution compared to traditional photoacoustic microscopy. Yang Fan and others from Shenzhen University have also made great progress in phase-type TIR ultrasonic sensing technology. The lateral resolution of the images obtained by the system they built has reached 5.8μm. Compared with traditional photoacoustic imaging systems, this system has higher sensitivity and wider detection bandwidth.

[0004] Although the existing phase-type TIR ultrasonic sensing system can already image tissue pathology well, this technology still has defects, such as the long distance between tissue samples and optical surface wave sensors, and the long distance that the photoacoustic high-frequency signal emitted by the samples needs to propagate in water. Figure 1As shown, tissue sample a and water layer b are placed on a glass slide c, and the glass slide c is placed on a two-dimensional displacement platform d. A prism e and an objective lens f are arranged above the two-dimensional displacement platform d. After the excitation light beam is focused by the objective lens f, it is incident on the prism e and then on the tissue sample a. The detection light beam emitted by the TIR detector g passes through the prism e and then is incident on the surface of the water layer b and is totally reflected. When the surface of the water layer b is close to the bottom surface of the prism e (that is, the distance between the prism e and the water layer b is minimized, and the movement of the two-dimensional displacement platform d is not interfered), according to the measurement data, the lower surface of the prism e will be 5 mm away from the tissue sample a at this time, and this 5 mm distance difference will have a great impact on the high-frequency information, that is, this will cause its acoustic wave frequency bandwidth to be reduced from 1 GHz to 170 MHz, which will directly lead to a decrease in the longitudinal resolution of the image and affect the three-dimensional imaging capability of the system. Summary of the invention

[0005] The purpose of the present invention is to provide a label-free photoacoustic pathology microscopic imaging system and imaging method, aiming to solve the problems of low longitudinal resolution of images generated by existing phase-type TIR ultrasonic sensing systems and poor coaxiality of existing detection beams.

[0006] In order to solve the above technical problems, the object of the present invention is achieved through the following technical solutions: providing a label-free photoacoustic pathology microscopy imaging system, which comprises:

[0007] An optical surface wave sensor, used for carrying a tissue sample and a water layer, wherein the tissue sample is immersed in the water layer;

[0008] An excitation light generating device, used for generating an excitation light beam;

[0009] A detection light generating device, used for generating a detection light beam;

[0010] an objective lens, used for focusing the excitation light beam and for converting the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam are incident on the tissue sample through the optical surface wave sensor;

[0011] A processing and control device for controlling the movement of the excitation light beam to scan the tissue sample;

[0012] The optical surface wave sensor is also used to totally reflect the detection light beam so that the reflected detection light beam is emitted through the objective lens;

[0013] The processing and control device is used to receive the returned detection light beam, and perform beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image.

[0014] In addition, the technical problem to be solved by the present invention is to provide an imaging method of a label-free photoacoustic pathology microscopy imaging system, which uses the label-free photoacoustic pathology microscopy imaging system as described above, and comprises:

[0015] placing a tissue sample and a water layer on the optical surface wave sensor, wherein the tissue sample is immersed in the water layer;

[0016] The excitation light generating device outputs an excitation light beam, and the detection light generating device outputs a detection light beam;

[0017] The objective lens focuses the excitation light beam and transforms the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam pass through the optical surface wave sensor and are incident on the tissue sample;

[0018] The processing and control device controls the movement of the excitation light beam to scan the tissue sample;

[0019] The optical surface wave sensor totally reflects the detection light beam, so that the reflected detection light beam is emitted through the objective lens;

[0020] The processing and control device receives the returned detection light beam, and performs beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image.

[0021] The embodiment of the present invention discloses a label-free photoacoustic pathology microscopic imaging system and an imaging method. The imaging system includes: an optical surface wave sensor, which is used to carry a tissue sample and a water layer, wherein the tissue sample is immersed in the water layer; an excitation light generating device, which is used to generate an excitation light beam; a detection light generating device, which is used to generate a detection light beam; an objective lens, which is used to focus the excitation light beam and to convert the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam are incident on the tissue sample through the optical surface wave sensor; a processing and control device, which is used to control the movement of the excitation light beam to scan the tissue sample; the optical surface wave sensor is also used to totally reflect the detection light beam, so that the reflected detection light beam is emitted through the objective lens; the processing and control device is used to receive the returned detection light beam, and perform beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image.

[0022] Compared with the prior art, the imaging system has a higher degree of integration and unification, which simplifies the complexity of the imaging system and makes the imaging system of the present application easier to apply in the field of biomedical imaging. Secondly, the system uses an optical surface wave sensor as a slide to carry tissue samples, so that the photoacoustic high-frequency signal emitted by the tissue sample does not need to propagate over long distances in water, and can retain the high-frequency signal to a great extent, thereby improving the longitudinal resolution, thereby realizing high-sensitivity, broadband photoacoustic detection. Thirdly, the imaging system of the present application allows the excitation light beam and the detection light beam to share an objective lens, so the coaxiality of the two light beams can be better guaranteed, thereby improving the lateral resolution of the imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings required for use in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other accompanying drawings can be obtained based on these accompanying drawings without paying any creative work.

[0024] Figure 1 It is a structural schematic diagram of an existing phase-type TIR ultrasonic sensing system;

[0025] Figure 2 A schematic diagram of the structure of a label-free photoacoustic pathology microscopy imaging system provided by an embodiment of the present invention;

[0026] Figure 3 A schematic flow chart of an imaging method of a label-free photoacoustic pathology microscopy imaging system provided in an embodiment of the present invention.

[0027] Description of reference numerals:

[0028] a, tissue sample; b, water layer; c, glass slide; d, two-dimensional displacement platform; e, prism; f, objective lens; g, TIR detector;

[0029] 1. Optical surface wave sensor; 2. Excitation light generating device; 3. Detection light generating device; 4. Objective lens; 5. Processing terminal; 6. Optical galvanometer; 7. Fifth reflector; 8. First lens; 9. Beam splitter; 10. First reflector; 11. Second reflector; 12. First analyzer; 13. Second analyzer; 14. Third reflector; 15. Fourth reflector; 16. Differential detector; 17. Second lens; 18. Sixth reflector; 19. Polarizer; 20. 1 / 2 wave plate; 21. 1 / 4 wave plate. DETAILED DESCRIPTION

[0030] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0031] It should be understood that when used in this specification and the appended claims, the terms "include" and "comprises" indicate the presence of described features, integers, steps, operations, elements and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components and / or combinations thereof.

[0032] It should also be understood that the terms used in this specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. As used in the specification of the present invention and the appended claims, unless the context clearly indicates otherwise, the singular forms "a", "an" and "the" are intended to include plural forms.

[0033] It should be further understood that the term "and / or" used in the present description and the appended claims refers to any and all possible combinations of one or more of the associated listed items, and includes these combinations.

[0034] See also Figure 2 , Figure 2 A schematic diagram of the structure of a label-free photoacoustic pathology microscopy imaging system provided by an embodiment of the present invention;

[0035] The label-free photoacoustic pathology microscopy system provided by the embodiment of the present invention comprises:

[0036] An optical surface wave sensor 1, used for carrying a tissue sample and a water layer, wherein the tissue sample is immersed in the water layer;

[0037] An excitation light generating device 2, used for generating an excitation light beam;

[0038] A detection light generating device 3, used for generating a detection light beam;

[0039] an objective lens 4, used for focusing the excitation light beam and for converting the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam are incident on the tissue sample through the optical surface wave sensor 1;

[0040] A processing and control device for controlling the movement of the excitation light beam to scan the tissue sample;

[0041] The optical surface wave sensor 1 is also used for totally reflecting the detection beam so that the reflected detection beam is emitted through the objective lens 4;

[0042] The processing and control device is used to receive the returned detection light beam, and perform beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image.

[0043] In this embodiment, firstly, a tissue sample (tissue or cell) and a water layer are placed directly on the optical surface wave sensor 1, an excitation light generating device 2 outputs an excitation light beam, a detection light generating device 3 outputs a detection light beam, an objective lens 4 focuses the excitation light beam, and at the same time, the objective lens 4 converts the detection light beam into a parallel light beam and emits it, and under the control of the processing and control device, the emitted excitation light beam and detection light beam are emitted to the tissue sample, wherein the excitation light beam and the detection light beam are incident on the tissue sample through the optical surface wave sensor 1, and the excitation light beam is driven to move by the control of the processing and control device to scan the tissue sample, in this embodiment, the detection light beam is reflected to the objective lens 4 after total reflection by the optical surface wave sensor 1, the processing and control device receives the detection light beam emitted after passing through the objective lens 4, and performs beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image, wherein the optical surface wave sensor 1 is used for total internal reflection of the return light beam to perform photoacoustic sensing.

[0044] The present application utilizes an optical surface wave sensor 1 to replace the role of the glass slide in the prior art, so that the optical surface wave sensor 1 can not only realize the role of carrying tissue samples, but also greatly reduce the distance between the optical surface wave sensor 1 and the tissue samples, so that the photoacoustic high-frequency signal emitted by the tissue sample does not need to propagate over long distances in the water, and the high-frequency signal can be retained to a great extent, thereby improving the longitudinal resolution.

[0045] It should be noted that in the prior art Figure 1 As shown, the lower surface of prism e is 5 mm away from tissue sample a at this time, which will cause the refractive index of water and the refractive index of prism e to not match, that is, it will cause aberration, reduce the focal light intensity, and cause the lateral resolution of the image to decrease. In the present application, since the tissue sample is directly placed on the optical surface wave sensor 1, the distance between the two is extremely small, and the adverse effects of the mismatch of the refractive index of water can be ignored. At the same time, in the prior art, since the excitation light beam and the detection light beam converge at different positions, they cannot be perfectly coaxial, which will also affect the imaging quality. Therefore, in the present application, the excitation light beam and the detection light beam are concentrated on the same objective lens 4, that is, since the two beams of light pass through the same objective lens 4, this can better ensure the coaxiality of the two beams of light, which can not only improve the lateral resolution of the imaging, but also ensure high-sensitivity photoacoustic signal detection, and can achieve accurate imaging of tissue pathology and even cell pathology.

[0046] In addition, the microscope imaging system of the present application simplifies the complexity of the system while achieving the imaging advantages of "label-free, high resolution, and high sensitivity", making the system more integrated.

[0047] In a specific embodiment, the objective lens 4 is located below the optical surface wave sensor 1 , and the optical galvanometer 6 is located below the objective lens 4 .

[0048] Through such a design, the complexity of the system can be simplified and the system structures can be made more compact.

[0049] In a specific embodiment, the excitation light generating device 2 is a laser with a wavelength of 266 nm, 532 nm or 560 nm, and the detection light generating device 3 is a helium-neon laser with a wavelength of 632.8 nm.

[0050] In this embodiment, 560nm excitation light is commonly used for hemoglobin imaging, 532nm excitation light is commonly used for melanin imaging, and 266nm excitation light is the optimal wavelength for cell nucleus imaging. That is to say, for different samples, the wavelength can be changed, based on the maximum absorption coefficient of the tissue sample and the strongest photoacoustic signal generated. By the same token, the wavelength of the detection light beam is not limited to 632.8nm, and different detection light wavelengths correspond to different excitation angles, so this application will not go into details.

[0051] In a specific embodiment, the processing and control device includes a processing terminal 5 and a plurality of optical galvanometers 6 connected to the processing terminal 5 by signal, wherein the processing terminal 5 is used to control the movement of the optical galvanometers 6, wherein the optical galvanometers 6 are located between the excitation light generating device 2 and the objective lens 4.

[0052] In this embodiment, the processing terminal 5 includes but is not limited to a computer. By inputting a movement instruction on the processing terminal 5, the movement of the optical galvanometer 6 can be controlled. In particular, the optical galvanometer 6 of the present application is provided with two. Compared with the phenomenon that the two-dimensional mobile platform of the prior art moves slowly, the present application can control the detection light beam to quickly and stably scan the tissue sample through the coordinated use of the two optical galvanometers 6 and the processing terminal 5. Compared with the existing photoacoustic imaging systems that mostly use a precision displacement control platform to move the tissue sample, the present application controls the movement of the optical galvanometer 6 through the processing terminal 5, which can overcome the problem that the slow movement speed of the two-dimensional displacement platform leads to a long scanning time and slow imaging speed.

[0053] In a specific embodiment, a plurality of reflectors and lenses are sequentially arranged between the optical galvanometer 6 and the excitation light generating device 2 along the light path direction, wherein the lens is used to expand the excitation light beam.

[0054] In this embodiment, the reflector is Figure 2 The fifth reflector 7 shown in the figure has one reflector 7 and is used to reflect the light beam and change the propagation direction of the light beam. Figure 2 As shown, two first lenses 8 are provided, both of which are used to expand the excitation light beam to ensure that the excitation light beam can fill the optical galvanometer 6. The present application does not specifically limit the number of the fifth reflector 7 and the first lens 8.

[0055] In a specific embodiment, a plurality of reflectors, polarizers 19 and wave plates are sequentially arranged between the detection light generating device 3 and the objective lens 4 along the light path direction.

[0056] In this embodiment, the reflector is Figure 2 The sixth reflector 18 shown is used to reflect the light beam and change the propagation direction of the light beam. The polarizer 19 is used to increase the linear polarization ratio of the detection light beam, and the wave plate is used to adjust the polarization direction of the linear polarized light.

[0057] In a specific embodiment, one of the reflector and polarizer is provided, and two of the wave plates are provided, and the two wave plates include a 1 / 2 wave plate 20 and a 1 / 4 wave plate 21 which are sequentially arranged along the light path direction.

[0058] In this embodiment, the 1 / 2 wave plate 20 is used to continuously adjust the polarization direction of the linear polarized light, and the 1 / 4 wave plate 21 is used to modulate the linear polarized light into elliptically polarized light. It should be noted that the initial detection light beam is not limited to linear polarized light, but can also be elliptically or circularly polarized light, whichever has the highest signal sensitivity.

[0059] In a specific embodiment, the processing and control device further includes a beam splitter 9, a first reflector 10, a second reflector 11, a first analyzer 12, a second analyzer 13, a third reflector 14, a fourth reflector 15, and a differential detector 16. The beam splitter 9 is located between the objective lens 4 and the terminal. The beam splitter 9 is used to split the returned detection light beam to obtain a first split beam and a second split beam.

[0060] The first reflector 10, the first analyzer 12, and the second reflector 11 are sequentially located in the optical path direction of the first split beam;

[0061] The third reflector 14, the second analyzer 13, and the fourth reflector 15 are sequentially located in the optical path direction of the second split beam;

[0062] The differential detector 16 is used to receive the first beam and the second beam, and perform photoacoustic signal synthesis;

[0063] The processing terminal 5 is signal-connected to the differential detector 16 and is also used to receive the synthesized photoacoustic signal and perform data analysis and image reconstruction.

[0064] In this embodiment, the beam splitting device 9 is a beam splitter, and the beam splitter is used to split the detection beam into two parts. In addition, the device for decomposing the mixed beam into s-polarized light and p-polarized light is not only a polarization beam splitter (PBS), but also a polarization beam splitter device such as a Wollaston prism.

[0065] The first reflector 10, the second reflector 11, the third reflector 14 and the fourth reflector 15 are used to reflect the light beam and change the propagation direction of the light beam. The first polarizer 12 and the second polarizer 13 are used to make the deflection direction of the polarized light propagate along the short axis direction or the long axis direction. The differential detector 16 is used to detect the photoacoustic signal.

[0066] In a specific embodiment, a plurality of lenses are arranged between the objective lens 4 and the beam splitting device 9 .

[0067] In this embodiment, the lens Figure 2 The second lens 17 shown in the figure has one lens and is used to focus the totally reflected detection light beam.

[0068] In this embodiment, 266nm excitation light in the microscopic imaging system is emitted by the excitation light generating device 2. After the excitation light beam propagates forward through the reflector 7, it is expanded by the 4f system composed of a pair of confocal lenses to ensure that the excitation light beam can fill the optical galvanometer 6. The optical galvanometer 6 is controlled by a computer connection to enable the expanded excitation light beam to perform fast and stable scanning on the tissue sample. The excitation light beam is focused onto the tissue sample through the objective lens 4. The tissue sample will emit a photoacoustic pulse wave under the action of the excitation light beam. The photoacoustic pulse wave will cause the refractive index of water to change. The photoacoustic wave propagates in the coupling medium water to the surface (above) of the optical surface wave sensor 1 and interacts with the evanescent field, wherein the evanescent field is generated when the detection light beam undergoes total internal reflection.

[0069] For the detection part, the system uses a 632.8nm laser (3) as a detection light source. After the emitted linear polarized light (detection beam) is reflected by the reflector, it passes through a polarizer to improve the polarization degree of the linear polarized light. The polarized light then passes through a 1 / 2 wave plate and a 1 / 4 wave plate, thereby modulating into elliptically polarized light with different phase differences in the s and p directions. After the elliptically polarized light passes through an objective lens 4, it will obtain parallel light that is irradiated on the optical surface wave sensor 1 for total internal reflection. Since the photoacoustic pulse wave obtained by the excitation light changes the refractive index of water, it will also change the phase difference of the reflected s and p polarized lights. That is to say, when the excitation light beam acts on the tissue sample, the tissue sample emits ultrasonic information. When the detection light beam passes through the interface between the optical surface wave sensor 1 and water, it will receive this information (that is, the phase change). Finally, the required image information is obtained by detecting the phase change.

[0070] The specific receiving process is as follows: the reflected light (detection beam) is focused by the second lens 17 and then passes through a beam splitter to divide the detection beam into two partial beams. The two beams are transmitted to two polarizers (12, 13) respectively. The deflection directions of the two are respectively along the short axis direction and the long axis direction of the elliptically polarized beam. The efficiency of converting the phase difference of the s and p beams caused by the acoustic wave in the short axis direction into light intensity is high, and it is mainly used for sensing. The conversion efficiency in the long axis direction is low, and it is used as a reference arm. After being deflected by the reflectors (10, 11, 14, 15), the two detection beams enter the two detection units of the differential detector 16 in parallel, that is, the differential detector 16 finally realizes the detection of the photoacoustic signal, and the photoacoustic signal is stored in a computer for subsequent data analysis and image reconstruction.

[0071] In the microscope imaging system provided by the present application, the optical surface wave sensor 1 is cleverly utilized, which not only plays the role of a carrier, but also can become the optical surface wave sensor 1 to transmit optical surface waves, so that the distance between the pathological tissue and the detection beam is closer, and the detection beam can better obtain high-frequency signals, thereby further improving the longitudinal resolution of the system imaging, and significantly simplifying the complexity of the system, making the system tend to be integrated, and easier to apply in the field of biomedical research.

[0072] The excitation light beam and the detection light beam of the present application are transmitted simultaneously by an objective lens 4, which can better ensure the coaxiality of the two light beams, improve the lateral resolution of imaging, ensure high-sensitivity photoacoustic signal detection, and make the system more miniaturized and improve integration.

[0073] Compared with traditional photoacoustic imaging systems, the microscopic imaging system of the present application has the characteristics of wider detection bandwidth, higher system sensitivity, and the fact that photoacoustic imaging does not require fluorescent labeling, providing better support for the "label-free, high-resolution, and high-sensitivity" observation of tissue samples.

[0074] In addition, Figure 3 As shown, the present invention also provides an imaging method of a label-free photoacoustic pathology microscopy imaging system, using the label-free photoacoustic pathology microscopy imaging system as described above, which comprises the following steps:

[0075] S101, placing a tissue sample and a water layer on the optical surface wave sensor 1, wherein the tissue sample is immersed in the water layer;

[0076] S102, the excitation light generating device 2 outputs an excitation light beam, and the detection light generating device 3 outputs a detection light beam;

[0077] S103, the objective lens 4 focuses the excitation light beam and transforms the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam pass through the optical surface wave sensor 1 and are incident on the tissue sample;

[0078] S104, the processing and control device controls the movement of the excitation light beam to scan the tissue sample;

[0079] S105, the optical surface wave sensor 1 totally reflects the detection light beam, so that the reflected detection light beam is emitted through the objective lens 4;

[0080] S106, the processing and control device receives the returned detection light beam, and performs beam splitting, photoacoustic signal synthesis and image reconstruction on the detection light beam to obtain a target image.

[0081] In this method, the optical surface wave sensor 1 is cleverly utilized, which not only plays the role of a carrier, but also can become the optical surface wave sensor 1 to transmit the optical surface wave, so that the distance between the pathological tissue and the detection beam is closer, that is, the detection beam can better obtain the high-frequency signal, thereby further improving the longitudinal resolution of the imaging.

[0082] The excitation light beam and the detection light beam of the present application are transmitted simultaneously by an objective lens 4, which can better ensure the coaxiality of the two light beams, improve the lateral resolution of imaging, and ensure high-sensitivity photoacoustic signal detection. At the same time, the system composed of the method of the present application is more miniaturized and the integration is improved.

[0083] Compared with the imaging method of the traditional photoacoustic imaging system, the imaging method of the microscopic imaging system of the present application has the characteristics of wider detection bandwidth, higher system sensitivity, and the characteristic that photoacoustic imaging does not require fluorescent labeling, providing better support for the "label-free, high-resolution, high-sensitivity" observation of tissue samples.

[0084] Those skilled in the art can clearly understand that, for the convenience and brevity of description, the specific working process of the method described above can refer to the corresponding process in the aforementioned system embodiment, and will not be repeated here.

[0085] The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can easily think of various equivalent modifications or replacements within the technical scope disclosed by the present invention, and these modifications or replacements should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be based on the protection scope of the claims.

Claims

1. A label-free photoacoustic pathology microscopy system, characterized in that: include: An optical surface wave sensor, used for carrying a tissue sample and a water layer, wherein the tissue sample is immersed in the water layer; An excitation light generating device, used for generating an excitation light beam; A detection light generating device, used for generating a detection light beam; an objective lens, used for focusing the excitation light beam and for converting the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam are incident on the tissue sample through the optical surface wave sensor; A processing and control device for controlling the movement of the excitation light beam to scan the tissue sample; The optical surface wave sensor is also used to totally reflect the detection light beam so that the reflected detection light beam is emitted through the objective lens; The processing and control device is used to receive the returned detection light beam, and perform beam splitting, photoacoustic signal synthesis and image reconstruction on the detection light beam to obtain a target image; The processing and control device comprises a processing terminal and a plurality of optical galvanometers connected to the processing terminal by signal, wherein the processing terminal is used to control the movement of the optical galvanometers, wherein the optical galvanometers are located between the excitation light generating device and the objective lens; The processing and control device further comprises a beam splitter, a first reflector, a second reflector, a first analyzer, a second analyzer, a third reflector, a fourth reflector, and a differential detector, wherein the beam splitter is located between the objective lens and the terminal, and is used to split the returned detection light beam to obtain a first split beam and a second split beam; The first reflector, the first analyzer, and the second reflector are sequentially located in the optical path direction of the first split beam; The third reflector, the second analyzer, and the fourth reflector are sequentially located in the optical path direction of the second split beam; The differential detector is used to receive the first beam splitter and the second beam splitter and perform photoacoustic signal synthesis; The processing terminal is connected to the differential detector signal and is also used to receive the synthesized photoacoustic signal and perform data analysis and image reconstruction.

2. The label-free photoacoustic pathology microscopy system according to claim 1, characterized in that: A plurality of reflectors and lenses are sequentially arranged between the optical galvanometer and the excitation light generating device along the light path direction, wherein the lens is used to expand the excitation light beam.

3. The label-free photoacoustic pathology microscopy system according to claim 1, characterized in that: A plurality of lenses are arranged between the objective lens and the beam splitting device.

4. The label-free photoacoustic pathology microscopy system according to claim 1, characterized in that: The objective lens is located below the optical surface wave sensor, and the optical galvanometer is located below the objective lens.

5. The label-free photoacoustic pathology microscopy system according to claim 1, characterized in that: The excitation light generating device is a laser with a wavelength of 266nm, 532nm or 560nm, and the detection light generating device is a helium-neon laser with a wavelength of 632.8nm.

6. The label-free photoacoustic pathology microscopy system according to claim 1, characterized in that: A plurality of reflectors, polarizing plates and wave plates are arranged in sequence between the detection light generating device and the objective lens along the light path direction.

7. The label-free photoacoustic pathology microscopy system according to claim 6, characterized in that: The reflector and polarizer are provided with one, the wave plates are provided with two, and the two wave plates include a 1 / 2 wave plate and a 1 / 4 wave plate which are sequentially arranged along the light path direction.

8. A label-free photoacoustic pathology microscopy method, using the label-free photoacoustic pathology microscopy system according to any one of claims 1 to 7, characterized in that: include: placing a tissue sample and a water layer on the optical surface wave sensor, wherein the tissue sample is immersed in the water layer; The excitation light generating device outputs an excitation light beam, and the detection light generating device outputs a detection light beam; The objective lens focuses the excitation light beam and transforms the detection light beam into a parallel detection light beam, so that the excitation light beam and the detection light beam pass through the optical surface wave sensor and are incident on the tissue sample; The processing and control device controls the movement of the excitation light beam to scan the tissue sample; The optical surface wave sensor totally reflects the detection light beam, so that the reflected detection light beam is emitted through the objective lens; The processing and control device receives the returned detection light beam, and performs beam splitting, photoacoustic signal synthesis and image reconstruction processing on the detection light beam to obtain a target image.

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