Multi-photon optical fiber array endoscopic imaging device and method based on hollow spiral scanning

Abstract

The invention discloses a multi-photon optical fiber array endoscopic imaging device and method based on hollow spiral scanning, and belongs to the technical field of biological imaging. The device comprises a laser collimation and beam expansion module, a pulse chirp module, a light beam switching module, an endoscopic detection module and a fluorescence acquisition module. The light beam switching module comprises an optical switch and an optical fiber array composed of at least two low-group-velocity dispersion single-mode optical fibers. The endoscopic detection module drives the optical fiber array to execute hollow spiral scanning through the resonance driver, the track starting point is not located at the view field original point, and central dense sampling is avoided. According to the method, the optical fibers are switched through the optical switch, so that the optical fibers execute spatial non-coincident hollow spiral scanning, multi-photon signals such as fluorescence, second harmonic and fluorescence lifetime are acquired, a central hole is complemented through image reconstruction and fusion, and a complete field imaging result is obtained. Light damage of the central area is remarkably reduced, the imaging resolution uniformity is improved, the fused image is true and reliable based on physical sampling, and the system stability is high.

Description

Technical Field

[0001] This invention relates to the field of bioimaging technology, and in particular to an endoscopic imaging device and method based on a hollow spiral scanning multiphoton fiber array. Background Technology

[0002] Currently, multiphoton microendoscopy technology has shown great application potential in the biomedical field due to its advantages such as no need for external labeling, strong tissue penetration, high sensitivity and specificity, excellent spatial resolution, and inherent optical slicing capabilities. Among them, multiphoton endoscopy systems based on piezoelectric ceramic tubes, gradient refractive index lenses (GRINlens), and low group velocity dispersion fibers have become the mainstream solution. These systems achieve helical scanning to complete imaging by driving a cantilevered fiber through a piezoelectric ceramic tube.

[0003] However, traditional helical scanning has significant drawbacks: the trajectory diameter increases over time, but the number of sampling points per revolution remains constant, resulting in a "dense in the center and sparse at the edges" distribution of samples. This characteristic leads to two major problems: first, the central region is prone to optical damage due to high-density repeated sampling, which is particularly serious in high-repetition-rate sampling scenarios such as multiphoton fluorescence lifetime imaging; second, insufficient sampling density in the edge regions leads to uneven image resolution and affects image quality.

[0004] To address the above problems, existing technologies mainly employ three types of solutions: Changing the scanning method: such as using grid scanning or Lissajous scanning, but this requires replacing system components (such as multi-core optical fibers and galvanometers), which can easily introduce new problems (such as resolution limitations due to the number of cores in multi-core optical fibers, and resource waste caused by the mismatch between Lissajous trajectories and circular fields of view), making it difficult to substantially improve system performance.

[0005] Optimizing spiral scanning parameters, such as hybrid spiral scanning and constant linear velocity spiral scanning, can improve resolution uniformity, but they are still based on the traditional spiral principle, and the sampling in the central region is still dense, which cannot fundamentally reduce the risk of light damage; some schemes (such as double spiral scanning) even increase the sampling density in the center.

[0006] Image post-processing algorithms: These algorithms improve image quality through deep learning and other methods, but may introduce pseudo-structures that reduce credibility. They also consume high computational resources, have poor interpretability, and are difficult to deploy in real time in small systems.

[0007] Therefore, there is an urgent need for a multiphoton endoscopic imaging technology that can reduce central light damage and improve resolution uniformity in principle, without requiring complex post-processing. Summary of the Invention

[0008] In view of the serious optical damage and non-uniform image resolution of traditional spiral scanning in the prior art, as well as the defects of existing optimization schemes, this invention aims to provide an endoscopic imaging device and method based on hollow spiral scanning multiphoton fiber array to achieve full field-of-view imaging with low optical damage and high resolution uniformity.

[0009] To achieve the above objectives, the present invention provides an endoscopic imaging device based on a hollow spiral scanning multiphoton fiber array, comprising: The laser collimation and beam expansion module is used to collimate and expand the femtosecond pulse laser emitted from the laser. The pulse chirp module, connected to the laser collimation and beam expander module, is used to pre-compensate for pulse broadening of the optical pulse after transmission through the optical fiber; The beam switching module, connected to the pulse chirp module, includes an optical switch and an optical fiber array. The optical fiber array includes at least two low group velocity dispersion single-mode fibers. The optical switch is used to selectively couple the laser after pulse chirp compensation to the target fiber in the optical fiber array. The endoscopic detection module, connected to the beam switching module, includes a resonant driver, a package housing, and a micro-objective. The resonant driver is used to drive one end of the fiber array to generate scanning motion. The package housing seals the end of the fiber array, the resonant driver, and the micro-objective. The micro-objective is used to focus the excitation light onto the sample and couple the multiphoton signal of the sample to the fiber array. The fluorescence acquisition module, connected to the beam switching module and the endoscopic detection module, is used to acquire multiphoton signals transmitted by the fiber optic array, perform image reconstruction and fusion processing, and obtain imaging results with a complete field of view. The resonant driver drives the fiber array to execute a hollow spiral scanning trajectory, the starting point of which is not located at the origin of the scanning field of view.

[0010] Preferably, the laser collimation and beam expansion module includes a laser, a first lens, and a second lens. The first lens and the second lens are sequentially arranged in the output optical path of the laser to collimate and expand the spatial light emitted from the laser.

[0011] Preferably, the pulse chirping module includes a beam splitter, a prism pair, and a reflector. The prism pair consists of a first prism and a second prism, and is used in conjunction with the reflector to compress the laser pulse to compensate for the broadening of the optical pulse during transmission through the optical fiber.

[0012] Preferably, the beam switching module further includes a second reflector, an optical fiber coupler, and a low group velocity dispersion fiber. The second reflector is used to reflect the compressed laser to the fluorescence acquisition module, and the optical fiber coupler is used to couple the spatial light to the low group velocity dispersion fiber. One end of the low group velocity dispersion fiber is connected to the optical fiber coupler, and the other end is connected to the input port of the optical switch.

[0013] Preferably, the fluorescence acquisition module includes a dichroic mirror, a bandpass filter group, a lens group, a photomultiplier tube group, a time-correlated single-photon counter, and a computer; the dichroic mirror includes a first dichroic mirror and a second dichroic mirror, used to separate the fluorescence signal and the second harmonic signal, respectively; the bandpass filter group includes a first bandpass filter and a second bandpass filter, used to filter signals in specific wavelength bands, respectively; the lens group includes a third lens and a fourth lens, used to focus the signal, respectively; the photomultiplier tube group includes a first photomultiplier tube and a second photomultiplier tube, used to acquire the fluorescence signal and the second harmonic signal, respectively; the time-correlated single-photon counter is used to acquire the fluorescence lifetime signal; and the computer is used to control image reconstruction and fusion processing. Among them, the third reflector is set in the optical path of the second harmonic signal and is used to refract the second harmonic signal to the second bandpass filter and the fourth lens.

[0014] This invention also provides a multiphoton fiber array endoscopic imaging method based on hollow spiral scanning, comprising the following steps: S1. Provide an optical fiber array, which includes at least two low group velocity dispersive single-mode fibers, and one end of the optical fiber array is driven by a resonant driver. S2. Control the laser to emit femtosecond pulse laser, which, after collimation, beam expansion, and pulse chirp compensation, is selectively coupled to the target fiber in the fiber array through the beam switching module; S3. Drive one end of the fiber array through a resonant driver to make the target fiber perform a hollow spiral scanning trajectory. The starting point of the hollow spiral scanning trajectory is not located at the origin of the scanning field of view. S4. Acquire multiphoton signals of the sample through an optical fiber array. The multiphoton signals include fluorescence signals and / or second harmonic signals. S5. Switch the beam switching module to other optical fibers in the fiber array, and repeat steps S2-S4 to make the other optical fibers perform hollow spiral scanning trajectories that do not overlap with the target optical fiber space. S6. Reconstruct images from the multiphoton signals collected by each optical fiber to obtain multiple sub-images; S7. Perform fusion processing on multiple sub-images to fill in the central hole region of each hollow spiral scanning trajectory and obtain the imaging result of the complete field of view.

[0015] Preferably, in step S3, the function of the hollow spiral scanning trajectory is: ; in, It is the radius from the current point to the origin. It's the angle. The initial radius and , The radial growth factor is the initial radius. The value ranges from 0.1 mm to 2 mm.

[0016] Preferably, in step S4, the multiphoton signal also includes a fluorescence lifetime signal, which is acquired by a time-correlated single-photon counter, and the laser provides a synchronization signal for the acquisition of the fluorescence lifetime signal.

[0017] Preferably, the resonant frequency of the resonant driver satisfy: ; in, It is the cantilever length. It is the Young's modulus of the material. It is the moment of inertia of the cross section. It is the material density. It is the cross-sectional area.

[0018] Therefore, the multiphoton fiber array endoscopic imaging device and method based on hollow spiral scanning with the above-described structure of the present invention has the following beneficial effects: (1) Reduce the risk of light damage: This invention avoids dense sampling in the central region by designing a hollow spiral scanning trajectory (initial radius a≠0), which significantly reduces the accumulation of light in the center and is especially suitable for light-sensitive samples and long-term imaging scenarios. (2) Improve resolution uniformity: In the fiber array of the present invention, multiple optical fibers perform complementary hollow spiral trajectories, and fill the central hole through image fusion to solve the problem of sparse edge sampling and achieve uniform sampling across the entire field of view. (3) The images are real and reliable: all pixels of the fused images of the present invention are derived from physical scanning sampling, avoiding the introduction of pseudo-structures by post-processing algorithms, and improving the credibility and clinical applicability of the results; (4) High system stability: The present invention uses optical switches to switch fiber channels, which simplifies the complex control of multi-core fibers, avoids crosstalk between cores, and improves system reliability; (5) Multimodal imaging capability: The present invention simultaneously acquires fluorescence intensity, fluorescence lifetime and second harmonic signal, enriches the dimensions of imaging information and meets diverse biological imaging needs.

[0019] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Attached Figure Description

[0020] Figure 1 This is a schematic diagram of the multiphoton fiber array endoscopic imaging device based on hollow spiral scanning according to the present invention. Figure 2This is a schematic diagram of the hollow spiral scanning sampling point distribution of the dual-core fiber array of the present invention; wherein, (a) and (c) are the sampling point distribution of the three sets of hollow spiral trajectory of fiber one, (d) is the sampling point distribution of the hollow spiral trajectory of fiber two, (e) is the sampling point distribution after fusion, and (f) is the sampling point distribution of traditional solid spiral scanning. Figure Labels 1-Laser, 2-First lens, 3-Second lens, 4-Beam splitter, 5-First prism, 6-Second prism, 7-Reflector 1, 8-Reflector 2, 9-Dichroic mirror 1, 10-Fiber coupler, 11-Low group velocity dispersive fiber, 12-Optical switch, 13-Fiber array, 14-Resonant driver, 15-Encapsulation housing, 16-Micro objective, 17-Sample, 18-Dichroic mirror 2, 19-Bandpass filter 1, 20-Third lens, 21-First photomultiplier tube, 22-Reflector 3, 23-Bandpass filter 2, 24-Fourth lens, 25-Second photomultiplier tube, 26-Controller, 27-Acquisition card, 28-Time-correlated single-photon counter, 29-Computer. Detailed Implementation

[0021] The technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments.

[0022] Unless otherwise defined, the technical or scientific terms used in this invention shall have the ordinary meaning understood by one of ordinary skill in the art to which this invention pertains. The terms "first," "second," and similar terms used in this invention do not indicate any order, quantity, or importance, but are merely used to distinguish different components. Terms such as "comprising" or "including" mean that the element or object preceding the word encompasses the elements or objects listed following the word and their equivalents, without excluding other elements or objects. Terms such as "connected" or "linked" are not limited to physical or mechanical connections, but can include electrical connections, whether direct or indirect. Terms such as "upper," "lower," "left," and "right" are used only to indicate relative positional relationships; when the absolute position of the described object changes, the relative positional relationship may also change accordingly.

[0023] Example like Figure 1 As shown, the present invention provides an endoscopic imaging device based on a hollow spiral scanning multiphoton fiber array, as detailed below: Laser collimation and beam expansion module: Laser 1 emits femtosecond pulsed laser (center wavelength 800nm, repetition frequency 80MHz), which is collimated and expanded by the first lens 2 (focal length 10mm) and the second lens 3 (focal length 20mm) to expand the laser spot diameter to 5mm; Pulse chirp module: The beam splitter 4 splits the laser into two beams. One beam is reflected by a prism pair consisting of the first prism 5 and the second prism 6 (made of SF11) and a reflector 7, thereby achieving pulse compression (compensating for the pulse broadening caused by fiber optic transmission, compressing the pulse width to 100 fs). Beam switching module: Mirror 2 8 refracts the compressed laser to dichroic mirror 1 9 (transmits 800nm ​​excitation light and reflects 500-600nm fluorescence), and couples it to low group velocity dispersion fiber 11 (length 2m, dispersion coefficient <1ps / (nm・km)) via fiber coupler 10 (coupling efficiency >85%), and transmits it to optical switch 12 (1×4 ports, switching time <1ms); Fiber array 13 consists of 4 low group velocity dispersion single-mode fibers arranged in a square with a close-end spacing of 50μm, and the other end is connected to the 4 output ports of optical switch 12 respectively; Endoscopic detection module: The dense end of the fiber optic array 13 is driven by a resonant driver 14 (piezoelectric ceramic resonant driver), and the encapsulation shell 15 (made of 304 stainless steel) provides waterproof and insulating sealing; the micro-objective 16 (numerical aperture 0.5, focal length 1.8 mm) focuses the excitation light onto the sample 17 and couples the fluorescence signal back to the fiber optic array 13; the resonant frequency of the resonant driver 14 is: ; Among them, cantilever length Young's modulus of material Moment of inertia of cross section Material density Cross-sectional area The calculated frequency is .

[0024] Fluorescence acquisition module: Dichroic mirror 18 (reflects 500-600nm fluorescence, transmits 800nm ​​second harmonic) separates the signal; the fluorescence signal is focused onto the first photomultiplier tube 21 (model H10722-01) by bandpass filter 19 (center wavelength 550nm, bandwidth 50nm) and third lens 20 (focal length 25mm); the second harmonic signal is refracted by mirror 22 and focused onto the second photomultiplier tube 25 (model H10722-01) by bandpass filter 23 (center wavelength 400nm, bandwidth 20nm) and fourth lens 24 (focal length 25mm); the time-correlated single-photon counter 28 (model PicoHarp300) acquires the fluorescence lifetime signal; computer 29 drives resonant driver 14 through acquisition card 27 (model NIPCIe-6363) and controller 26, and runs image fusion algorithm.

[0025] Imaging process Taking a dual-core fiber array (fiber 1 and fiber 2) as an example, the imaging process is as follows: Laser 1 emits a femtosecond laser, which is collimated, expanded, and chirped before being transmitted to optical switch 12 via fiber coupler 10 and low group velocity dispersion fiber 1. Optical switch 12 opens the first fiber channel, and laser is coupled to fiber 1; resonant driver 14 drives fiber array 13, causing fiber 1 to execute a hollow spiral trajectory: , ( , The acquisition card 27 works synchronously with the time-correlated single-photon counter 28 to obtain... Figure 2 The sub-image of the sampling point distribution shown in (a) in the image; Adjust the scan parameters to The optical fiber was scanned again to obtain... Figure 2 The sub-image shown in (b) is shown in the image. Adjust the scan parameters to The fiber optic cable was scanned a third time to obtain... Figure 2 The sub-image shown in (c) is shown in the image. Optical switch 12 switches to fiber optic channel 2, fiber optic channel 2 execution trajectory Scan to obtain Figure 2 The sub-image shown in (d) is shown in the image. 29 pairs of computers Figure 2 (a) in Figure 2 The sub-images of (d) are fused: through spatial registration (based on sample feature point matching), bilinear interpolation is used to fill in the central holes to generate... Figure 2 The complete image shown in (e) is compared. Figure 2 Compared with the traditional solid spiral scanning result in (f), the present invention has no central dense sampling and the overall sampling is more uniform.

[0026] In summary, this invention is applicable to multiphoton fluorescence and fluorescence lifetime imaging systems. This method significantly reduces laser light damage in the central region of traditional spiral scanning trajectories by designing a hollow spiral scanning trajectory; and by employing multiple fibers in an fiber array to perform hollow spiral scanning with non-overlapping trajectories or sampling points, the resulting multiple reconstructed images are fused to achieve full-field multiphoton fluorescence and fluorescence lifetime imaging, while effectively improving the overall resolution uniformity of the imaging image.

[0027] This invention employs an optical switch as a switching device for the excitation beam, achieving efficient coupling between the excitation light and different optical fibers in the fiber array. Based on the hollow spiral scanning method described above, the system can achieve low-optical-damage imaging of samples. The image data acquired by each fiber in the fiber array is processed by reconstruction and fusion algorithms to obtain a complete field-of-view endoscopic imaging result. All pixels in the imaging result originate from physical sampling of the actual spiral scan, ensuring the authenticity and reliability of the image.

[0028] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. A multiphoton fiber optic array endoscopic imaging device based on hollow spiral scanning, characterized in that, include: The laser collimation and beam expansion module is used to collimate and expand the femtosecond pulse laser emitted from the laser. The pulse chirp module, connected to the laser collimation and beam expander module, is used to pre-compensate for pulse broadening of the optical pulse after transmission through the optical fiber; The beam switching module, connected to the pulse chirp module, includes an optical switch and an optical fiber array. The optical fiber array includes at least two low group velocity dispersion single-mode fibers. The optical switch is used to selectively couple the laser after pulse chirp compensation to the target fiber in the optical fiber array. The endoscopic detection module, connected to the beam switching module, includes a resonant driver, a package housing, and a micro-objective. The resonant driver is used to drive one end of the fiber array to generate scanning motion. The package housing seals the end of the fiber array, the resonant driver, and the micro-objective. The micro-objective is used to focus the excitation light onto the sample and couple the multiphoton signal of the sample to the fiber array. The fluorescence acquisition module, connected to the beam switching module and the endoscopic detection module, is used to acquire multiphoton signals transmitted by the fiber optic array, perform image reconstruction and fusion processing, and obtain imaging results with a complete field of view. The resonant driver drives the fiber array to execute a hollow spiral scanning trajectory, the starting point of which is not located at the origin of the scanning field of view.

2. The multiphoton fiber optic endoscopic imaging device based on hollow spiral scanning according to claim 1, characterized in that: The laser collimation and beam expansion module includes a laser, a first lens, and a second lens. The first lens and the second lens are sequentially arranged in the output optical path of the laser to collimate and expand the spatial light emitted from the laser.

3. The multiphoton fiber optic endoscopic imaging device based on hollow spiral scanning according to claim 1, characterized in that: The pulse chirp module includes a beam splitter, a prism pair, and a reflector. The prism pair consists of a first prism and a second prism, which are used in conjunction with the reflector to compress the laser pulse to compensate for the broadening of the optical pulse during transmission through the optical fiber.

4. The multiphoton fiber optic endoscopic imaging device based on hollow spiral scanning according to claim 1, characterized in that: The beam switching module also includes a second reflector, an optical fiber coupler, and a low group velocity dispersion fiber. The second reflector is used to reflect the compressed laser to the fluorescence acquisition module. The optical fiber coupler is used to couple the spatial light to the low group velocity dispersion fiber. One end of the low group velocity dispersion fiber is connected to the optical fiber coupler, and the other end is connected to the input port of the optical switch.

5. The multiphoton fiber optic endoscopic imaging device based on hollow spiral scanning according to claim 1, characterized in that: The fluorescence acquisition module includes a dichroic mirror, a bandpass filter group, a lens group, a photomultiplier tube group, a time-correlated single-photon counter, and a computer. The dichroic mirror includes dichroic mirror one and dichroic mirror two, used to separate fluorescence signals and second harmonic signals, respectively. The bandpass filter group includes bandpass filter one and bandpass filter two, used to filter signals in specific wavelength bands, respectively. The lens group includes a third lens and a fourth lens, used to focus signals, respectively. The photomultiplier tube group includes a first photomultiplier tube and a second photomultiplier tube, used to acquire fluorescence signals and second harmonic signals, respectively. The time-correlated single-photon counter is used to acquire fluorescence lifetime signals. The computer is used to control image reconstruction and fusion processing. Among them, the third reflector is set in the optical path of the second harmonic signal and is used to refract the second harmonic signal to the second bandpass filter and the fourth lens.

6. The multiphoton fiber optic endoscopic imaging device based on hollow spiral scanning according to claim 1, characterized in that: Resonant frequency of the resonant driver f satisfy: ； in, It is the cantilever length. It is the Young's modulus of the material. It is the moment of inertia of the cross section. It is the material density. It is the cross-sectional area.

7. A multiphoton fiber optic array endoscopic imaging method based on hollow spiral scanning, applied to the multiphoton fiber optic array endoscopic imaging device based on hollow spiral scanning as described in any one of claims 1-6, characterized in that, Includes the following steps: S1. Provide an optical fiber array, which includes at least two low group velocity dispersive single-mode fibers, and one end of the optical fiber array is driven by a resonant driver. S2. Control the laser to emit femtosecond pulse laser, which, after collimation, beam expansion, and pulse chirp compensation, is selectively coupled to the target fiber in the fiber array through the beam switching module; S3. Drive one end of the fiber array through a resonant driver to make the target fiber perform a hollow spiral scanning trajectory. The starting point of the hollow spiral scanning trajectory is not located at the origin of the scanning field of view. S4. Acquire multiphoton signals of the sample through an optical fiber array. The multiphoton signals include fluorescence signals and / or second harmonic signals. S5. Switch the beam switching module to other optical fibers in the fiber array, and repeat steps S2-S4 to make the other optical fibers perform hollow spiral scanning trajectories that do not overlap with the target optical fiber space. S6. Reconstruct images from the multiphoton signals collected by each optical fiber to obtain multiple sub-images; S7. Perform fusion processing on multiple sub-images to fill in the central hole region of each hollow spiral scanning trajectory and obtain the imaging result of the complete field of view.

8. The multiphoton fiber optic endoscopic imaging method based on hollow spiral scanning according to claim 7, characterized in that: In step S3, the function of the hollow spiral scanning trajectory is: ； in, It is the radius from the current point to the origin. It's the angle. The initial radius and , The radial growth factor is the initial radius. The value ranges from 0.1 mm to 2 mm.

9. The endoscopic imaging method based on hollow spiral scanning multiphoton fiber array according to claim 7, characterized in that: In step S4, the multiphoton signal also includes a fluorescence lifetime signal, which is acquired by a time-correlated single-photon counter, and the laser provides a synchronization signal for the acquisition of the fluorescence lifetime signal.

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