Multi-photon microscopic imaging probe, imaging device and imaging method

Abstract

The invention discloses a multi-photon microscopic imaging probe, imaging equipment and an imaging method. The imaging probe comprises a medium assembly, and the medium assembly comprises a solid elastic medium and an imaging window sheet which carries the solid elastic medium and is used for being in contact with an imaging object. And the medium assembly is detachably arranged at the first end of the handle-shaped shell. The imaging module is arranged in the handle-shaped shell, is connected with the photoelectric composite cable and is used for transmitting and controlling the exciting light and receiving the fluorescence signal, and the imaging module comprises an objective lens which is in contact fit with the solid elastic medium. Therefore, the objective lens provided by the invention is in contact with the imaging object based on the solid medium, and the limitation of a traditional immersed medium on a microscopic imaging device is eliminated, so that the imaging probe can be in contact with the imaging object at any angle.

Description

Technical Field

[0001] This specification relates to the field of microscopic imaging, and particularly to a multi-photon microscopic imaging probe, an imaging device, and an imaging method. Background Art

[0002] Multi-photon microscopy is a non-linear optical imaging technique based on the multi-photon absorption / excitation phenomenon. Among them, multi-photons generally include one or more imaging modalities such as two-photon, three-photon, second harmonic, and third harmonic. The multi-photon absorption / excitation phenomenon refers to a non-linear optical phenomenon involving the interaction of multiple photons with matter in an optical process.

[0003] In the application of label-free in-vivo imaging, since the autofluorescence of biological tissues is very weak, the imaging system is required to have extremely high imaging performance, stable imaging performance, and user-friendliness. Compared with traditional fluorescence imaging techniques, the wavelength of the excitation light used in multi-photon microscopy is in the near-infrared band, and microscopic imaging can be performed on the internal structure of the imaging object within a certain depth range. However, in actual imaging, due to problems such as the stability of the focal plane, environmental light interference, and internal scattering of the imaging object in in-vivo microscopy compared with traditional stage microscopy, the imaging quality will be seriously affected. Therefore, how to improve the quality of in-vivo multi-photon microscopy is a technical problem that needs to be solved urgently by those skilled in the art. Summary of the Invention

[0004] In view of this, the embodiments of this specification provide a multi-photon microscopic imaging probe, an imaging device, and an imaging method. The objective lens contacts the imaging object through a solid elastic medium, reducing the aberration between the medium and the imaging object, thereby overcoming the deficiencies of the prior art.

[0005] In a first aspect, the embodiments of this specification provide a multi-photon microscopic imaging probe, which includes: a medium assembly, where the medium assembly includes a solid elastic medium and an imaging window for carrying the solid elastic medium and contacting the imaging object. A handle-shaped housing, where the medium assembly is detachably disposed at the first end of the handle-shaped housing. And an imaging module based on micro-laser scanning microscopy technology, where the imaging module is disposed in the handle-shaped housing and is used to transmit and control the excitation light and receive the fluorescence signal, and the imaging module includes an objective lens in contact with the solid elastic medium.

[0006] In some embodiments of this specification, the imaging module further includes a displacement stage for controlling the overall movement of the imaging module within the handle-shaped housing to adjust the focal plane position of the imaging module.

[0007] In certain embodiments of this specification, the medium component further includes a first fixing structure, and a second fixing structure matching the first fixing structure is provided at the first end of the handle-shaped housing. The medium component is detachably disposed at the first end of the handle-shaped housing through the first fixing structure and the second fixing structure.

[0008] In certain embodiments of this specification, the first fixing structure is annularly disposed around the imaging window and extends along the placement direction of the solid elastic medium.

[0009] In certain embodiments of this specification, the first fixing structure is made of a light-shielding material. When the first fixing structure cooperates with the second fixing structure and the imaging window contacts the imaging object, a low-light environment is formed between the imaging object and the objective lens.

[0010] In certain embodiments of this specification, the refractive index of the solid elastic medium matches the imaging object.

[0011] In certain embodiments of this specification, the elastic deformation range of the solid elastic medium matches the imaging depth range.

[0012] In certain embodiments of this specification, the imaging probe further includes an optoelectronic composite cable for connecting to an imaging host. The optoelectronic composite cable is connected to the second end of the handle-shaped housing and is connected to the imaging module. The optoelectronic composite cable includes a first optical fiber for transmitting excitation light, a second optical fiber for transmitting fluorescence signals, and a control bus for transmitting control signals.

[0013] In a second aspect, an embodiment of this specification provides a multi-photon microscopy imaging device, which includes: an imaging host for generating excitation light and generating a fluorescence image based on the fluorescence signal; and the multi-photon microscopy imaging probe as shown in the first aspect. Among them, the multi-photon microscopy imaging probe is connected to the imaging host through an optoelectronic composite cable, and is used to release excitation light to an imaging object and collect the fluorescence signal generated by the imaging object based on the photoluminescence effect.

[0014] In a third aspect, an embodiment of this specification provides a multi-photon microscopy imaging method, which is applied to the multi-photon microscopy imaging probe as shown in the first aspect or the multi-photon microscopy imaging device as shown in the second aspect. The method includes: controlling the position of the imaging module in the imaging direction so that the focal plane of the imaging module is at a target focusing depth. Among them, when the focal plane is at the target focusing depth, the objective lens is in contact and cooperation with the solid elastic medium. Performing multi-photon microscopy imaging on the imaging object based on the target focusing depth.

[0015] The embodiments of this specification provide a multi - photon microscopy imaging probe, an imaging device, and an imaging method. By using a solid elastic medium as the medium for indirect contact between the objective lens and the imaging object, the use limitations of traditional immersion media on the microscopy imaging device are eliminated. As a result, the imaging probe can contact the imaging object at any angle and can perform reciprocating imaging in the depth dimension. In addition, in this specification, the solid elastic medium is cured into a medium component and connected to the probe body (handle - shaped housing) in a detachable form. During use, the assembly of the objective lens and the solid elastic medium can be completed directly by disassembling the medium component, without the frequent and complex assembly process of traditional immersion media. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] To more clearly illustrate the technical solutions of the embodiments of the present invention, the following will briefly introduce the drawings required for use in the embodiments. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as limiting the scope. For those of ordinary skill in the art, without creative efforts, other related drawings can also be obtained based on these drawings.

[0017] Figure 1 It is a schematic diagram of the imaging situation based on the contact between the immersion medium and the imaging object provided by an exemplary embodiment of this specification.

[0018] Figure 2 It is an application scenario diagram of a multi - photon microscopy imaging device provided by an exemplary embodiment of this specification.

[0019] Figure 3 It is a schematic structural diagram of a multi - photon microscopy imaging probe provided by an exemplary embodiment of this specification.

[0020] Figure 4 It is a schematic diagram showing the detachable setting of the medium component and the handle - shaped housing provided by an exemplary embodiment of this specification.

[0021] Figure 5 It is a schematic diagram of the contact - type cooperation between the objective lens and the solid elastic medium provided by an exemplary embodiment of this specification.

[0022] Figure 6 It is an exemplary flowchart of a multi - photon microscopy imaging device provided by an exemplary embodiment of this specification.

[0023] Figure 7 It is an exemplary flowchart of a multi - photon microscopy imaging method provided by an exemplary embodiment of this specification.

[0024] Figure 8 It is an exemplary flowchart of a method for determining a solid elastic medium provided by an exemplary embodiment of this specification.

[0025] Among them, 110 is an optical element; 120 is an imaging sample; 130 is an immersion environment; 140 is an immersion medium; 10 is an imaging device; 20 is an imaging object; 200 is an imaging probe; 300 is an imaging host; 400 is an optoelectronic composite cable; 221 is a solid elastic medium; 222 is an imaging window; 231 is an objective lens; 230 is an imaging module; 220 is a medium component; 210 is a handle-shaped housing; 232 is a displacement stage; 223 is a first fixing structure; 211 is a second fixing structure; 310 is a power supply; 320 is a laser light source; 330 is a collection module; 340 is a signal control and processing module. Detailed implementation manners

[0026] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present application. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present application without creative efforts shall fall within the protection scope of the present application.

[0027] Application Overview:

[0028] During the multi-photon microscopy imaging process, the influence of the imaging object on the excitation light is mainly manifested as aberration. Among them, aberration refers to the deviation of the imaging quality during the imaging process due to the refraction and reflection of light when passing through different parts of the optical system, resulting in image distortion or blurring. Different refractive indices will cause light to refract when entering and passing through different media, thereby affecting the propagation direction and speed of light and resulting in aberration.

[0029] Exemplarily, the refractive index of human skin generally ranges between 1.3 and 1.6, while the refractive index of the traditional air medium is 1. This means that when the excitation light enters the human skin from the air, obvious refraction will occur, resulting in aberration.

[0030] To reduce aberration, in some existing multi-photon microscopy devices, an immersion medium is used instead of the air medium, thereby reducing the aberration between the medium and the imaging object. For example, water or a similar light-transmitting liquid mixture can be used as the immersion medium. Exemplarily, the refractive index of water is 1.33, and the refractive index can be changed by mixing with specific reagents or using specific liquid media, so that the refractive index is close to that of human skin. Thus, when the excitation light enters the human skin from the liquid immersion medium, no obvious refraction will occur and the aberration is small.

[0031] Based on the immersion medium, when performing multi-photon microscopy imaging, there are generally two implementation methods: complete immersion and partial immersion.

[0032] Figure 1 This is a schematic diagram of an imaging situation provided by an exemplary embodiment of this specification, based on the contact between an immersion medium and an imaging object.

[0033] As Figure 1 shown, the schematic diagram of the imaging situation may specifically include a full-immersion mating relationship 100a based on an immersion environment and a partial-immersion mating relationship 100b based on an immersion slide. In each schematic of the mating relationship, the description is based on the optical element 110 and the imaging sample 120. Among them, the optical element 110 only represents a part of the optical elements (such as an objective lens) that emit excitation light in the existing embodiments, and can also be understood as a collection of optical devices in a multi-photon microscopy device. The imaging sample 120 can be an imaging object that meets the corresponding restrictions in various mating relationships. For example, an ex vivo sample.

[0034] In the full-immersion mating relationship 100a, the immersion medium can form an immersion environment 130, so that the optical element 110 and the imaging sample 120 are fully immersed in the immersion environment 130.

[0035] As Figure 1 shown, during imaging, both the optical element 110 and the imaging sample 120 are placed inside the immersion environment 130, and the imaging direction of the optical element 110 is aligned with the imaging sample 120. Based on this, the excitation light released by the optical element 110 enters the inside of the imaging sample 120 through the immersion environment 130. When the refractive index of the immersion environment is close to that of the imaging sample 120, there will be no obvious refraction phenomenon when the excitation light enters the imaging object, and the aberration is small.

[0036] Based on the above technical solution, when performing multi-photon microscopy imaging, an immersion environment must be formed, which thus limits the imaging object (generally an ex vivo object) and the imaging scene (which must be immersed in the immersion environment).

[0037] In the schematic diagram of the partial-immersion mating relationship 100b, the immersion medium 140 can build an immersion slide through a cover glass, so that the objective lens is immersed in the immersion environment on the immersion slide.

[0038] As Figure 1 shown, during imaging, the cover glass can carry the liquid immersion medium 140, and the liquid immersion medium forms a small spherical-like immersion environment due to its own surface tension. The cover glass is in contact with the imaging sample 120, and the optical element 110 is immersed in the immersion environment formed by the immersion medium 140.

[0039] Thus, the excitation light released by the optical element 110 always enters the imaging sample 120 through the immersion medium 140 and the cover glass to form a focal plane. During depth imaging, the position of the focal plane within the imaging sample 120 can be adjusted by adjusting the position of the objective lens. Among them, when the refractive index of the immersion medium 140 matches that of the imaging sample 120, the displacement of the optical element 110 is consistent with the displacement of the focal plane. During the entire imaging process, the refractive index on the path of the excitation light forming the focal plane remains basically unchanged. Thus, the aberration is greatly reduced.

[0040] Based on the above technical solutions, there are at least the following technical problems at the practical application level:

[0041] First, the immersion medium 140 itself is affected by gravity, which limits the application scenarios of the imaging device. Specifically, the immersion medium 140 itself is a fluid or semi-fluid. When the cover glass is placed in a non-horizontal direction, the immersion medium 140 itself will be displaced under the action of gravity, so that the aforementioned small immersion environment cannot be stably formed, and thus imaging cannot be performed. For example, when the immersion medium 140 is coated on the skin, it will slide down quickly. In addition, when the immersion medium 140 is in a zero-gravity environment, the immersion medium 140 cannot achieve stable contact with the objective lens and the imaging window, so imaging cannot be performed.

[0042] Second, the immersion medium 140 is a disposable device based on its own properties, that is, after the imaging task is completed, the shape of the immersion medium 140 cannot be restored and cannot be reused, and the immersion slide often needs to be reassembled. Exemplarily, when the optical element 110 moves in the immersion medium 140, affected by the volume of the optical element 110 immersed in the immersion medium 140, the lateral area of the immersion medium 140 will expand significantly. When the optical element 110 leaves the immersion medium 140, its shape is quite different from the original shape and cannot be reused to perform subsequent imaging tasks and needs to be reassembled.

[0043] In the related art, the immersion medium may also include a colloidal medium. Based on the adsorption cooperation between the colloidal medium and the imaging window and the objective lens, some of the above technical problems can be overcome. However, in practice, the colloidal medium still cannot completely solve the above technical problems. Exemplarily, for the colloidal medium, when the objective lens moves towards the imaging window, the colloidal medium can achieve adsorption cooperation with the imaging window and the objective lens. However, when the objective lens moves away from the imaging window, the colloidal medium may be adsorbed to the imaging window and the objective lens respectively, splitting the colloidal medium into two parts, resulting in gaseous medium mixed in the middle, and thus generating aberration. Based on this situation, it can be seen that the colloidal medium is still a disposable device and needs to be reassembled each time it is used. For imaging at special angles, it is still affected by gravity and there is a possibility of inability to image.

[0044] Based on the above technical problems, the embodiments of the present application provide an imaging probe and an imaging device based on a solid elastic medium. By using a solid elastic medium as the medium for indirect contact between the objective lens and the imaging object, the use limitations of traditional immersion media on the microscopic imaging device are eliminated, so that the imaging device can contact the imaging object at any angle in the form of an imaging probe and can reciprocally image at the depth level. In addition, in this specification, the solid elastic medium is solidified into a medium component and connected to the probe body (handle-shaped housing) in a detachable form. During use, the assembly of the objective lens and the solid elastic medium can be completed directly by disassembling the medium component, without the frequent and complex assembly process of traditional immersion media. The various non-limiting embodiments of the present application will be specifically introduced below with reference to the accompanying drawings.

[0045] Exemplary Application Scenarios:

[0046] Figure 2 FIG. 1 is an application scenario diagram of a multi-photon microscopy device provided by an exemplary embodiment of this specification.

[0047] As Figure 2 shown, the application scenario diagram of the multi-photon microscopy device may include a multi-photon microscopy device 10 and an imaging object 20. The multi-photon microscopy device 10 may be based on a solid elastic medium and adopt a split design, and thus further includes a multi-photon microscopy probe 200 and an imaging host 300. Among them, the multi-photon microscopy probe 200 may cooperate with the imaging object 20 based on a solid elastic medium, and the multi-photon microscopy probe 200 and the imaging host 300 may be communicatively connected based on an optoelectronic composite cable 400. Among them, more content about the multi-photon microscopy device 10 can be found in Figure 5 and its related descriptions.

[0048] As Figure 2 shown in the partial schematic diagram, the area where the multi-photon microscopy probe 200 contacts the imaging object 20 may include a solid elastic medium 221, an imaging window 222 for carrying the solid elastic medium, and an objective lens 231. Among them, more content about the multi-photon microscopy probe can be found in Figure 3 and its related descriptions.

[0049] When the multi - photon microscopy probe 200 performs multi - photon microscopy, the objective lens 231 is in contact - fit with the solid elastic medium 221. One side of the imaging window 222 bears the solid elastic medium 221, and the other side is in contact - fit with the imaging object 20. At this time, the excitation light generated by the objective lens 231 enters the interior of the imaging object 20 through the solid elastic medium 221 and the imaging window 222. When the objective lens 231 (imaging module 230) is moved, the solid elastic medium 221 compresses or stretches due to its own elastic properties, thus ensuring that the objective lens 231 remains in contact - fit with the solid elastic medium 221. Based on the fact that the solid elastic medium 221 is in a contact - fit state (compressed state) during imaging, the elastic force generated by its compressed state can be used to overcome the influence of gravity, so that the imaging probe 200 can contact the imaging object 20 at any angle. In addition, in a zero - gravity environment, the solid elastic medium 221 can also be fixed due to its own elastic force, so that multi - photon microscopy can still be performed in a zero - gravity environment. Among them, for the contact - fit between the objective lens 231 and the solid elastic medium 221, reference can be made to Figure 4 and its related descriptions.

[0050] Exemplary Imaging Probes:

[0051] Based on the foregoing application scenarios, this specification provides a multi - photon microscopy probe based on a solid elastic medium. Figure 3 is a schematic structural diagram of a multi - photon microscopy probe provided by an exemplary embodiment of this specification.

[0052] As Figure 3 shown, the multi - photon microscopy probe 200 may include an optoelectronic composite cable 400, a handle - shaped housing 210, a medium assembly 220, and an imaging module 230.

[0053] The optoelectronic composite cable 400 can be used to connect to the imaging host 300 and realize data transmission between the imaging host 300 and the imaging probe 200. Among them, the optoelectronic composite cable 400 can transmit the excitation light generated by the imaging host 300 into the imaging probe 200 and transmit the fluorescence signal collected by the imaging probe 200 to the imaging host 300 to form a fluorescence image.

[0054] In some embodiments, each device in the imaging probe 200 can be connected to the imaging host 300 based on the optoelectronic composite cable 400, so as to perform multi - photon microscopy under the control of the imaging host 300. In some embodiments, the optoelectronic composite cable 400 may include a first optical fiber for transmitting excitation light, a second optical fiber for transmitting fluorescence signals, and a control bus for transmitting control signals. Among them, the control bus can be an integrated control cable for each device in the imaging probe 200.

[0055] The handle-shaped housing 210 can be the outer shell of the multi-photon microscopy probe 200, and the device integration of the multi-photon microscopy probe 200 is arranged inside the handle-shaped housing 210. Among them, the handle-shaped housing 210 can be understood that relevant staff can hold the handle-shaped housing 210 by a holding method similar to holding a handle.

[0056] As Figure 3 shown, the handle-shaped housing 210 can be presented as a quasi-cylindrical structure, and is provided with a second end and a first end. Among them, the quasi-cylindrical structure between the second end and the first end forms a holding part. The second end is used to access the fiber optic composite cable 400 so that the fiber optic composite cable 400 is correspondingly connected to the imaging module 230. The first end is used for detachably cooperating with the medium component 220, and is used for when the medium component 220 is arranged at the first end, contacting the imaging object 20 through the imaging window 222 in the medium component 220. The holding part is used to be held by relevant staff / robotic arm, so as to contact the imaging object 20 at a preset angle.

[0057] In some embodiments, an opening can be provided at the first end of the handle-shaped housing 210, and the objective lens 231 is arranged at the opening of the first end. When the medium component 220 is assembled at the first end, the solid elastic medium 221 can be in contact and cooperate with the objective lens 231. Exemplarily, when the medium component 220 is assembled at the first end, the axis of the solid elastic medium 221 coincides with the axis of the objective lens 231, so that the solid elastic medium 221 can be in contact and cooperate with the objective lens 231 by displacing the objective lens 231. For more content about the cooperation relationship between the solid elastic medium 221 and the objective lens 231, reference can be made to Figure 4 and its related description.

[0058] The medium component 220 can refer to a component that realizes indirect contact between the objective lens 231 and the imaging object 20. Among them, the medium component 220 can be realized based on the aforementioned solid elastic medium 221 and the imaging window 222. That is, the medium component 220 can include the solid elastic medium 221 and the imaging window 222. One side of the imaging window 222 bears the solid elastic medium 221 and is fixedly connected to the solid elastic medium 221, and the other side is used for contact and cooperation with the imaging object 20 when performing multi-photon microscopy.

[0059] In some embodiments, the medium component 220 can be detachably arranged at the first end of the handle-shaped housing 210. The detachable arrangement can refer to realizing the detachable cooperation (fixed connection state and separation state) between the medium component 220 and the second end through mechanical components. For more content about the detachable cooperation, reference can be made to Figure 4 and its related description.

[0060] The solid elastic medium 221 can be a solid, elastic material with good light transmittance. In some embodiments, when the solid elastic medium 221 is in contact fit with the objective lens 231, the solid elastic medium 221 itself is compressed by the pressure of the objective lens 231. When the objective lens 231 moves further, the solid elastic medium 221 can cooperate through compression and stretching, so as to ensure the contact fit between the objective lens 231 and the solid elastic medium 221. In some embodiments, the elastic modulus of the solid elastic medium 221 can be related to the imaging depth range. For specific details, please refer to Figure 5 and its related content.

[0061] In some embodiments, the refractive index of the solid elastic medium 221 can be matched with the imaging object 20. Herein, the matching may mean that the refractive index of the solid elastic medium 221 is within the allowable range of the refractive index of the imaging object 20 (such as ±5%, ±10%). The specific material of the solid elastic medium 221 can be selected based on the imaging object 20. For example, if the refractive index of the skin of group A people is generally 1.45, then silicone gel (the refractive index can be made about 1.45 by doping), butyl rubber (refractive index about 1.46), etc. can be selected as the solid elastic medium. Another example is that if the refractive index of the skin of group B people is generally above 1.3, then fluororubber (refractive index 1.35) can be selected as the solid elastic medium.

[0062] It should be noted that in actual use, the refractive index of the solid elastic medium 221 may not be exactly the same as that of the imaging object 20. It can also be used only based on the characteristic that the solid elastic medium 221 resists gravity by elasticity during use, that is, the imaging probe 200 is used at different imaging angles.

[0063] The imaging window 222 can refer to the medium that directly contacts the imaging object 20. The working principle of the imaging window 222 is the same as that of the cover glass in a microscope sample, so as to facilitate cleaning the side that contacts the imaging object 20. The imaging window 222 can adopt common cover glass materials, such as glass.

[0064] In some embodiments, the imaging window 222 can also be integrally arranged in the solid elastic medium 221. For example, the side of the solid elastic medium 221 close to the imaging object 20 can be cured to form the imaging window 222.

[0065] In some embodiments, to further reduce aberration, the refractive index of the imaging window 222 can also be matched with that of the imaging object 20. Among them, the imaging window 222 can also be selected based on the refractive index of the imaging object 20. That is, the imaging window 222 can be selected as a solid light-transmitting medium with a refractive index matching that of the imaging object 20 and capable of forming a smooth interface. For example, for the skin of population A, the imaging window 222 can be selected as ultra-clear glass (refractive index 1.45 to 1.55).

[0066] The imaging module 230 can be understood as a collection of optical elements within the imaging probe 200. Among them, the imaging module 230 can construct the internal optical path of the imaging probe 200 based on the micro-laser scanning microscopy technology, thereby realizing the control of the excitation light.

[0067] The inside of the imaging module 230 can include an excitation light optical path and a fluorescence signal optical path. Among them, the excitation light optical path controls the excitation light to perform point-by-point scanning on the imaging object 20, and the fluorescence signal optical path is used to collect the fluorescence signal generated after the imaging object 20 is excited. Among them, the excitation light optical path can be connected to the first optical fiber in the optoelectronic composite cable 400, and the fluorescence signal optical path can be connected to the second optical fiber in the optoelectronic composite cable 400.

[0068] In some embodiments, the internal optical path of the imaging module 230 can be constructed based on the micro-laser scanning microscopy technology. For example, reference can be made to the internal structure schematic diagram shown in CN109662696A. Figure 7 In addition, the internal optical path of the imaging module 230 can be adjusted according to actual needs. For example, a collimation module can be set at the interface where the first optical fiber accesses the imaging module 230 for collimating the excitation light.

[0069] The objective lens 231 can refer to the optical element in indirect contact with the imaging object 20 in the imaging module 230, and can serve as the window for the excitation light optical path and the fluorescence signal optical path to contact the imaging object 20, for releasing the excitation light and collecting the fluorescence signal.

[0070] When the excitation light transmitted in the optoelectronic composite cable 400 is controlled by the imaging module 230 and emitted from the objective lens 231, the processed excitation light will form a stable focal plane. Among them, each emitted excitation light will include multiple photons, and the multiple photons are focused at a focal position on the focal plane, thereby activating the imaging object 20 corresponding to the focal position. The imaging module 230 can control the focal position of the excitation light through internal optical devices (such as a scanning galvanometer) to scan the imaging object 20 at the focal plane.

[0071] Considering that the relative position between the focal plane and the objective lens 231 is fixed, the focal plane position within the imaging object 20 can be adjusted by adjusting the position of the objective lens 231, thereby realizing the imaging of the internal structure of the imaging object 20.

[0072] In some embodiments, considering that the imaging module 230 itself is based on micro-laser scanning microscopy technology, when adjusting the position of the objective lens 231 alone, it is difficult for other internal optical elements to cooperate in the adjustment. Then, the imaging module 230 can be moved as a whole within the handle-shaped housing 210 to adjust the focal plane position. That is, the imaging module 230 further includes a displacement stage 232. Among them, the displacement stage 232 is used to control the overall movement of the imaging module 230 within the handle-shaped housing 210 to adjust the focal plane position of the imaging module 230.

[0073] In some embodiments, each component inside the imaging module 230 can be controlled based on the control bus in the optoelectronic composite cable 400. Among them, the control bus can specifically include a position control cable for controlling the displacement stage, a scanning control cable for controlling the scanning galvanometer, a communication cable for communicating with the imaging probe 200 body (such as the internal integrated circuit and control board of the imaging probe 200), and a ground wire.

[0074] Based on the multi-photon microscopy imaging probe provided in some embodiments of this specification, by using a solid elastic medium as the medium for indirect contact between the objective lens and the imaging object, the use limitations of traditional immersion media for microscopy imaging devices are eliminated, so that the imaging probe can contact the imaging object at any angle and can image reciprocally in the depth dimension. In addition, in this specification, the solid elastic medium is cured into a medium component and connected to the probe body (handle-shaped housing) in a detachable form. When in use, the assembly of the objective lens and the solid elastic medium can be completed directly by disassembling the medium component, without the frequent and complex assembly process of traditional immersion media.

[0075] Exemplary Detachable Fit:

[0076] Figure 4 It is a schematic diagram of the detachable setting of the medium component and the handle-shaped housing provided in an exemplary embodiment of this specification.

[0077] As Figure 4 shown, the detachable fit can include a separated state 400a and a fixed connection state 400b. When the medium component 220 and the handle-shaped housing 210 are in the separated state 400a, the two are separated. When the medium component 220 and the handle-shaped housing 210 are in the fixed connection state 400b, the axis of the solid elastic medium 221 coincides with the axis of the objective lens 231, and the objective lens 231 can be moved to make contact fit between the objective lens 231 and the solid elastic medium 221.

[0078] As Figure 4As shown, the medium component 220 may further include a first fixing structure 223 for realizing a detachable connection. A second fixing structure 211 matching the first fixing structure 223 is provided at the first end of the handle-shaped housing 210. Wherein, the medium component 220 may be detachably disposed at the first end of the handle-shaped housing 210 through the first fixing structure 223 and the second fixing structure 211.

[0079] In some embodiments, the first fixing structure 223 and its corresponding second fixing structure 211 may be common detachable components. For example, the first fixing structure 223 may be a card slot or a groove, and the second fixing structure 211 may be a pin for being held on the card slot or the groove. For another example, the first fixing structure 223 and the second fixing structure 211 may both be threads, wherein the thread ridges and thread valleys of the first fixing structure 223 and the second fixing structure 211 correspond to each other.

[0080] In some embodiments, to avoid affecting the contact between the imaging window 222 and the imaging object 20, the first fixing structure 223 may be disposed around the imaging window 222 and extend along the placement direction of the solid elastic medium 221. That is, the medium component 220 may form a concave structure, wherein the imaging window 222 is disposed on the bottom surface of the concave structure, the solid elastic medium 221 is disposed on the inner wall of the bottom surface, and the first fixing structure 223 is disposed on the side wall of the concave structure.

[0081] Corresponding to the first fixing structure 223, a boss structure corresponding to the concave structure may be formed at the first end of the handle-shaped housing 210, and the second fixing structure 211 may be disposed on the side wall of the boss structure, so that the concave structure and the boss structure cooperate to detachably dispose the medium component 220 at the first end.

[0082] Considering that the imaging probe 200 provided in this specification can directly contact the imaging object 20, imaging can be performed based on a low-light environment during imaging. Among them, low-light environment imaging may refer to blocking external light sources during the imaging process and using excited light and fluorescence signals as the main light sources for imaging.

[0083] In some embodiments, a low-light environment may be realized through the first fixing structure 223. That is, the first fixing structure 223 may be made of a light-shielding material. When the first fixing structure 223 cooperates with the second fixing structure 211 (i.e., the fixed connection state 400b) and the imaging window 222 contacts the imaging object 20 (i.e., the imaging window 222 does not introduce external light sources), a low-light environment is formed between the imaging object 20 and the objective lens 231.

[0084] Based on the first fixing structure 223 on the medium component 220 provided in some embodiments of this specification, not only can the detachable cooperation with the second fixing structure 211 be realized, but also external light sources can be blocked, thereby forming a low-light environment to improve the recognition effect of fluorescence signals.

[0085] Exemplary Contact Fit:

[0086] Figure 5 It is a schematic diagram of the contact cooperation between an objective lens and a solid elastic medium provided in an exemplary embodiment of this specification.

[0087] In some embodiments, the cooperation process between the objective lens and the solid elastic medium may include a non-contact state and a contact cooperation state. Among them, as Figure 5 shown in the example of contact cooperation, it may include a separation state 500a, a reference state 500b, an intermediate state 500c, and an extreme state 500d. Among them, the separation state 500a is a kind of non-contact state, and the reference state 500b, the intermediate state 500c, and the extreme state 500d are a kind of contact cooperation state.

[0088] The non-contact state may refer to a state where the contact cooperation between the objective lens 231 and the solid elastic medium 221 is not achieved. Specifically, it may include the separation state 500a.

[0089] When in the separation state 500a, the solid elastic medium 221 is separated from the objective lens 231. At this time, the solid elastic medium 221 is not compressed, and its height relative to the imaging window 222 can be denoted as h0. The objective lens 231 can form a focal plane. The focal plane can be seen in the trapezoidal area marked by a dotted line in the figure. Its bottom surface can be the focal plane, and the trapezoidal body can be regarded as the set of excitation lights forming the focal plane. Among them, when the excitation light is emitted from the objective lens 231, it will converge at the focal plane as shown in the figure. When in the non-contact state, the position of the focal plane generally does not reach the contact interface between the imaging window 222 and the imaging object 20.

[0090] When the objective lens 231 in the separation state moves towards the solid elastic medium 221, when the objective lens 231 and the solid elastic medium 221 meet the cooperation conditions, the objective lens 231 and the solid elastic medium 221 form a contact cooperation.

[0091] The cooperation conditions may include the following aspects of restrictions:

[0092] First, the objective lens 231 is in contact with the solid elastic medium 221, and the solid elastic medium 221 is deformed by force.

[0093] Second, the light rays forming the focal plane do not pass through media other than the solid elastic medium. That is, the excitation light forming the focal plane only passes through the solid elastic medium and the imaging window before entering the imaging object 20.

[0094] Third, the focal plane is located inside the imaging object 20.

[0095] In some embodiments, the contact area between the solid elastic medium 221 and the imaging window 222 is larger than the focal plane area. In this case, to ensure that the foregoing first and second conditions are met, it is only necessary to ensure that the contact interface formed by the objective lens 231 and the solid elastic medium 221 is horizontal with the focal plane and can completely cover the area where the excitation light is released in the objective lens 231.

[0096] Based on the above matching conditions, during the contact matching process in which the objective lens 231 moves towards the solid elastic medium 221, the reference state 500b, the intermediate state 500c, and the limit state 500d can be sequentially formed.

[0097] The reference state 500b can refer to the matching state when the focal plane is located at the position of the contact interface between the imaging window 222 and the imaging object 20.

[0098] At this time, the solid elastic medium 221 is compressed, and its height relative to the imaging window 222 can be denoted as h1. The relative depth between the focal plane of the objective lens 231 and the imaging object 20 is 0.

[0099] The intermediate state 500c can refer to the matching state when the focal plane is located inside the imaging object 20 and the objective lens 231 can reciprocate. Among them, the objective lens 231 can reciprocate can be understood as that in the moving direction of the objective lens 231, the objective lens 231 can move towards the imaging object 20 within a certain range and can also move away from the imaging object 20.

[0100] At this time, the solid elastic medium 221 is compressed, and its height relative to the imaging window 222 can be denoted as h2. The relative depth between the focal plane of the objective lens 231 and the imaging object 20 is d1.

[0101] The limit state 500d can refer to the matching state when the objective lens 231 cannot continue to move towards the imaging object 20 (i.e., the limit compression state of the solid elastic medium 221).

[0102] At this time, the solid elastic medium 221 is compressed to the limit value, and its height relative to the imaging window 222 can be denoted as h3. The relative depth between the focal plane of the objective lens 231 and the imaging object 20 is d2.

[0103] During actual imaging, the contact fit at least includes an intermediate state. Whether to include other states can be determined according to the actual imaging task. For example, when the imaging task includes detecting the relative depth from 0 to the target depth, it can first be in the reference state to determine the depth 0, then adjust the depth of the objective lens to the intermediate state to perform imaging, so that the relative depth between the focal plane and the imaging object 20 reaches the target depth. Another example is that when the imaging task includes the first target depth to the second target depth, it can directly be in the intermediate state to perform imaging, so that d1 is respectively the target depths from the first target depth to the second target depth.

[0104] It should be noted that considering that the refractive index of the solid elastic medium matches the imaging object, the displacement of the objective lens (or the deformation of the solid elastic medium) is basically the same as the displacement of the focal plane. For example, the height difference between the aforementioned h1 and h3 (i.e., the displacement distance of the objective lens from the reference state to the limit state) is the same as d2.

[0105] Based on the aforementioned fit relationship, it can be known that the elastic deformation range of the solid elastic medium can match the imaging depth range. Among them, the imaging depth range is the imaging depth range required by the imaging task. As shown in the aforementioned states, the elastic deformation range of the solid elastic medium can include the elastic range required for contact fit (from h0 to h1) and the elastic range after contact (from h1 to h3), and the imaging depth range can be any range from 0 to d2. Then it can be known that the elastic range after contact in the elastic deformation range of the solid elastic medium (such as the height difference from h1 to h3) is greater than or equal to the imaging depth range (generally any range from 0 to d2).

[0106] It should be noted that considering that different medium components may be replaced for the same probe facing different imaging tasks. The contact fit between the medium component and the objective lens only needs to meet the requirements of the imaging task. For example, when the position of the focal plane coincides with the contact interface between the imaging window and the imaging object, the objective lens and the solid elastic medium can be in a non-contact state. When implementing the contact fit subsequently, directly perform imaging on the imaging object within the imaging depth range based on the aforementioned intermediate state. Among them, the position of the focal plane can be directly determined according to the displacement parameters of the objective lens itself.

[0107] Exemplary Imaging Devices:

[0108] Figure 6 It is an exemplary flowchart of a multi-photon microscopy imaging device provided by an exemplary embodiment of this specification.

[0109] As Figure 6As shown, the multi - photon microscopy imaging device 10 may include an imaging host 300 and a multi - photon microscopy imaging probe 200. The imaging host 300 and the multi - photon microscopy imaging probe 200 are connected by an optical - electrical composite cable 400. Among them, for the specific internal structure of the multi - photon microscopy imaging probe 200, reference can be made to the relevant description in the foregoing Figure 3 , which will not be elaborated here.

[0110] The imaging host 300 may be a set of components in the imaging device 10 that cannot or are not convenient to be integrated into the imaging probe 200. For example, the imaging host 300 can be used to generate excitation light and generate a fluorescence image based on the fluorescence signal. The imaging host 300 is connected to the multi - photon microscopy imaging probe 200 through the optical - electrical composite cable 400 to transmit the excitation light and control signals and receive the fluorescence signal.

[0111] As Figure 6 shown, the imaging host 300 may specifically include a power supply 310, a laser light source 320, a collection module 330, and a signal control and processing module 340. Among them, the power supply 310 can be used to provide the working voltage for the imaging device 10. The laser light source 320 can be used to generate excitation light. The collection module 330 can be used to generate a fluorescence image based on the fluorescence signal. The signal control and processing module 340 is used to generate the control signal for controlling the imaging probe 200. The cable interface is used to connect to the optical - electrical composite cable 400.

[0112] Exemplary Imaging Methods:

[0113] Figure 7 is an exemplary flowchart of a multi - photon microscopy imaging method provided by an exemplary embodiment of this specification. In some embodiments, Figure 7 the process P700 shown can be executed by the signal control and processing module 340. Among them, when the signal control and processing module 340 executes P700, it needs to be realized by generating the corresponding control signal transmitted to the imaging probe 200. Therefore, P700 can also be understood as being executed by the imaging probe 200 or the imaging device 10.

[0114] As Figure 7 shown, P700 includes the following steps:

[0115] S710, control the position of the imaging module in the imaging direction so that the focal plane of the imaging module is at the target focusing depth. Among them, when the focal plane is at the target focusing depth, the objective lens is in contact and cooperation with the solid - state elastic medium.

[0116] The target focusing depth can refer to any value within the imaging depth range of an imaging task, which can reflect the position where the focal plane should be during imaging. For example, in a general imaging task, the imaging depth range can be from 0 to a preset depth. Exemplarily, when imaging the human facial skin, it is generally from 0 to 0.3 mm so that the imaging depth range includes all structures of the human epidermis.

[0117] In some embodiments, adjusting the focal plane to the target focusing depth is generally determined based on the displacement amount relative to the reference state. For example, the reference scale of the objective lens displacement in the reference state can be determined through image features, and then the target focusing depth can be reached by displacing the target focusing depth based on this reference scale. Among them, the reference state can also be determined based on the factory parameters of the objective lens and the medium component.

[0118] S720, perform multi-photon microscopy imaging on the imaging object based on the target focusing depth.

[0119] Considering that the focal plane is already at the target focusing depth, the scanning operation can be directly performed, thereby realizing multi-photon microscopy imaging at the target focusing depth. Only as an example, the focusing position of the excitation light in the focal plane can be adjusted by a scanning galvanometer, so that the focusing position traverses the entire focal plane. When the imaging object is excited by the excitation light at the corresponding position, a fluorescence signal can be generated, which is collected by the imaging probe and transmitted to the collection module. The collection module and the controller determine the fluorescence image at the target focusing depth based on the scanning timing signal and the fluorescence signal to complete the multi-photon microscopy imaging.

[0120] Exemplary Method for Determining Solid State Elastic Medium:

[0121] For the aforementioned imaging device based on a solid-state elastic medium, this application Figure 8 also provides a method for determining a solid-state elastic medium suitable for an imaging module based on an imaging task.

[0122] Figure 8 is an exemplary flowchart of the solid-state elastic medium determination method provided by some embodiments of this application.

[0123] As Figure 8 shown, P800 may include the following steps:

[0124] S810, determine the imaging device, imaging task, and candidate solid-state elastic media.

[0125] The imaging task can refer to at least one imaging content that the imaging device needs to perform during actual imaging. Among them, the imaging task can include the imaging object and the imaging depth.

[0126] In S810, the imaging device may refer to a device for performing actual imaging. In some embodiments, the imaging device may determine before performing the imaging task that the imaging task to be performed can be completed by the imaging device. In some embodiments, the imaging device may also be determined according to the imaging task. For example, an imaging device with better fluorescence effect can be selected according to the imaging object for imaging. For another example, an imaging device whose imaging module moving range can meet the imaging depth is selected according to the imaging depth.

[0127] The candidate solid-state elastic medium may refer to a solid-state elastic medium that can be selected in the current imaging task. That is, for the imaging task, the imaging device needs to determine a solid-state elastic medium that can perform the imaging task from the candidate solid-state media, and cooperate with the solid-state elastic medium to perform the imaging task. Among them, the candidate solid-state elastic medium can be directly determined from the existing elastic media.

[0128] In some embodiments, considering the imaging principle of the imaging device, the solid-state elastic medium can be screened to a certain extent to determine the candidate solid-state elastic medium, so as to reduce the workload of testing. Among them, the candidate solid-state elastic medium should at least satisfy that there is a cross-section larger than the light exit port / objective lens of the imaging module, and the imaging window corresponding to the candidate solid-state elastic medium is larger than the focal plane of the imaging module. Based on this, the determined candidate solid-state elastic medium can at least ensure that the candidate solid-state elastic medium and the imaging module can cooperate to have the aforementioned initial cooperation state.

[0129] S820, determine the reference position parameter, mating position parameter, and limit position parameter of the imaging module based on the candidate solid-state elastic medium.

[0130] The reference position parameter may refer to the position parameter (such as the position scale) of the displacement stage when in the reference state 500b in Figure 5 . The configuration position parameter may refer to the position parameter of the displacement stage when the objective lens and the solid-state elastic medium are just in contact and mating. The limit position parameter may refer to the position parameter of the displacement stage when in the limit state 500d in Figure 5 .

[0131] In some embodiments, the aforementioned mating position parameter can be determined based on the shape of the solid-state elastic medium. When the cross-sectional area of the solid-state elastic medium is larger than the light exit area of the objective lens, it can be considered that when the solid-state elastic medium deforms to this point, the objective lens and the solid-state elastic medium are just in contact and mating.

[0132] In some embodiments, the aforementioned parameters may also include the protruding height of the solid-state elastic medium relative to the imaging device. As Figure 4 shown, the solid-state elastic medium and the imaging window can be assembled on the imaging device. When assembling, there may be a certain distance between the top of the solid-state elastic medium and the end of the imaging device, and this distance can be denoted as the protruding height.

[0133] In some embodiments, the reference position parameter can be estimated based on the difference in the protruding height of different solid elastic media. For example, if the protruding height of solid elastic medium a is h1 and the reference position parameter is s1, for solid elastic medium b with a protruding height of h2, its reference position parameter can be s1 + h1 - h2.

[0134] In some embodiments, the protruding height of the candidate solid elastic medium can be marked after production and directly called during use. In some embodiments, the difference in the protruding height between different solid elastic media can also be determined based on testing, so as to determine the reference position parameter of each solid elastic medium.

[0135] In some embodiments, the mating position parameter can be determined based on the shape of the solid elastic medium. Among them, when the cross-sectional area of the solid elastic medium is larger than the light exit port of the imaging module (such as the area of the objective lens), it can be considered that the solid elastic medium covers the imaging module.

[0136] Similar to the aforementioned reference position parameter, in some embodiments, the mating position parameter can be determined based on the compression amount of the solid elastic medium when it is larger than the light exit port of the imaging module. For example, for solid elastic medium b, when its compression amount in the reference mating state is y1 and its compression amount in the initial mating state is y2, the mating position parameter of solid elastic medium b can be (s1 + h1 - h2) - y1 + y2.

[0137] In some embodiments, the limit position parameter can be determined according to the compression amount of the solid elastic medium in the limit compression state. The specific determination process is similar to that of the mating position parameter and will not be elaborated here.

[0138] S830, determine the target solid elastic medium from the candidate solid elastic media based on the reference position parameter, the mating position parameter, and the limit position parameter.

[0139] The target solid elastic medium can refer to a solid elastic medium that meets the requirements of the imaging task and can be used to perform subsequent imaging tasks.

[0140] In some embodiments, the imaging range corresponding to the solid elastic medium can be compared with the imaging depth to determine the target solid elastic medium. When the reference position parameter of the solid elastic medium is within the interval [mating position parameter, limit position parameter], the corresponding imaging range is the interval [0, limit position parameter - reference position parameter]. When the imaging depth of the imaging task is within this interval, it can be determined that the solid elastic medium meets the imaging task and can be used as the target solid elastic medium.

[0141] In some embodiments, when the imaging depth of the imaging task is relatively large, a combination of solid-state elastic media with multiple different imaging ranges can be used as the target solid-state elastic media.

[0142] When combining the solid-state elastic media, a solid-state elastic media with a reference position parameter < a matching position parameter can be considered. When the reference position parameter of the solid-state elastic media < the matching position parameter, the corresponding imaging range is the interval [matching position parameter - reference position parameter, limit position parameter - reference position parameter]. Since its reference position parameter is relatively small, its imaging range is relatively large. Although it does not include the 0 point, it can be combined with the solid-state elastic media that includes the 0 point, so as to cover the imaging depth.

[0143] Thus, the imaging device provided by the present application can select an appropriate solid-state elastic media based on the imaging task, thereby ensuring the smooth execution of the imaging task.

[0144] All of the above optional technical solutions can be combined arbitrarily to form optional embodiments of the present application, which will not be elaborated herein one by one.

[0145] Those of ordinary skill in the art can realize that the units and algorithm steps of each example described in combination with the embodiments disclosed herein can be implemented by electronic hardware, or a combination of computer software and electronic hardware. Whether these functions are executed in a hardware or software manner depends on the specific application and design constraints of the technical solution. Professional technicians can use different methods to implement the described functions for each specific application, but such implementation should not be considered to exceed the scope of the present application.

[0146] Those skilled in the art can clearly understand that for the convenience and brevity of description, the specific working processes of the systems, devices, and units described above can refer to the corresponding processes in the foregoing method embodiments, which will not be elaborated herein.

[0147] In several embodiments provided by the present application, it should be understood that the disclosed systems, devices, and methods can be implemented in other ways. For example, the device embodiments described above are merely illustrative. For example, the division of the units is only a logical function division. In actual implementation, there can be other division methods. For example, multiple units or components can be combined or integrated into another system, or some features can be ignored or not executed. Another point is that the displayed or discussed mutual coupling, direct coupling, or communication connection can be through some interfaces. The indirect coupling or communication connection of the devices or units can be in an electrical, mechanical, or other form.

[0148] The unit described as a separation component may or may not be physically separated. The component shown as a unit may or may not be a physical unit, that is, it may be located in one place or distributed over multiple network units. Some or all of the units can be selected according to actual needs to achieve the purpose of the solution of this embodiment.

[0149] In addition, in each embodiment of the present application, each functional unit may be integrated in a processing unit, may exist separately as individual physical units, or two or more units may be integrated in one unit.

[0150] If the described function is implemented in the form of a software functional unit and sold or used as an independent product, it can be stored in a computer-readable storage medium. Based on this understanding, the technical solution of the present application, in essence, or the part that contributes to the prior art or a part of this technical solution can be embodied in the form of a software product. This computer software product is stored in a storage medium and includes several instructions for causing a computer device (which may be a personal computer, a server, or a network device, etc.) to execute all or part of the steps of the methods described in each embodiment of the present application. The aforementioned storage medium includes: various media such as USB flash drives, mobile hard disks, read-only memories (ROM, Read-Only Memory), random access memories (RAM, Random Access Memory), magnetic disks, or optical discs that can store program verification codes.

[0151] It should be noted that in the description of the present application, the terms "first", "second", "third", etc. are only used for descriptive purposes and cannot be understood as indicating or implying relative importance. In addition, in the description of the present application, unless otherwise specified, the meaning of "a plurality of" is two or more.

[0152] The above are only the preferred embodiments of the present application and are not intended to limit the present application. Any modifications, equivalent replacements, etc. made within the spirit and principles of the present application shall be included within the protection scope of the present application.

Claims

1. A multiphoton microscopic imaging probe, characterized in that: The imaging probe comprises: A medium component, wherein the medium component comprises a solid elastic medium and an imaging window that carries the solid elastic medium and is used to contact an imaging object; a handle-shaped housing, wherein the media assembly is detachably disposed at a first end of the handle-shaped housing; and An imaging module based on micro laser scanning microscopy technology, wherein the imaging module is arranged in the handle-shaped shell and is used to transmit and control excitation light and receive fluorescence signals, and the imaging module includes an objective lens that is in contact with the solid elastic medium.

2. The imaging probe according to claim 1, characterized in that: The imaging module further comprises a displacement stage, and the displacement stage is used to control the overall movement of the imaging module in the handle-shaped housing to adjust the focal plane position of the imaging module.

3. The imaging probe according to claim 1, characterized in that: The media component also includes a first fixing structure, the first end of the handle-shaped shell is provided with a second fixing structure matching the first fixing structure, and the media component is detachably arranged on the first end of the handle-shaped shell through the first fixing structure and the second fixing structure.

4. The imaging probe according to claim 3, characterized in that: The first fixed structure is arranged around the imaging window and extends along the placement direction of the solid elastic medium.

5. The imaging probe according to claim 4, characterized in that: The first fixed structure is made of a light-shielding material. When the first fixed structure cooperates with the second fixed structure and the imaging window is in contact with the imaging object, a weak light environment is formed between the imaging object and the objective lens.

6. The imaging probe according to claim 1, characterized in that: The refractive index of the solid elastic medium matches the imaging object.

7. The imaging probe according to claim 1, characterized in that: The elastic deformation range of the solid elastic medium matches the imaging depth range.

8. The imaging probe according to claim 1, characterized in that: The imaging probe further comprises an optoelectronic composite cable for connecting to an imaging host, wherein the optoelectronic composite cable is connected to the second end of the handle-shaped housing and is connected to the imaging module; The optoelectronic composite cable includes a first optical fiber for transmitting excitation light, a second optical fiber for transmitting a fluorescent signal, and a control bus for transmitting a control signal.

9. A multiphoton microscopic imaging device, characterized in that: The device comprises: an imaging host for generating excitation light and generating a fluorescence image based on the fluorescence signal; and The multi-photon microscopic imaging probe according to any one of claims 1 to 8, wherein the multi-photon microscopic imaging probe is connected to the imaging host via an optoelectronic composite cable, and is used to release the excitation light to the imaging object and collect the fluorescence signal generated by the imaging object based on the photoluminescence effect.

10. A multiphoton microscopic imaging method, characterized in that: Applied to the multiphoton microscopic imaging probe according to any one of claims 1 to 8 or the multiphoton microscopic imaging device according to claim 9, the method comprises: Controlling the position of the imaging module in the imaging direction so that the focal plane of the imaging module is at a target focusing depth, wherein when the focal plane is at the target focusing depth, the objective lens is in contact with the solid elastic medium; Multiphoton microscopic imaging is performed on the imaging object based on the target focus depth.

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