A wide-field spinning-disk confocal tomographic microscopy system

Abstract

The utility model discloses a wide field of view turntable confocal chromatography microscopic imaging system, including shell and its inside installation laser light source, be provided with first relay lens module and dichroic mirror on the emergent light path of laser light source, be equipped with pinhole turntable and second relay lens module on the transmission light path of dichroic mirror, second relay lens module is located above sample, be provided with third relay lens module and detector on the reflection light path of dichroic mirror, second relay lens module and third relay lens module constitute conjugated optical module, second relay lens module is by two camera lenses common constitution imaging objective, third relay lens module is by two camera lens combination formation, the system still installs signal synchronous processing module, the utility model solves the problem that the turntable confocal microscopic system expands field of view usually leads to spatial resolution to drop, and pursues high resolution and has to shrink the problem of imaging field of view.

Description

Technical Field

[0001] This utility model relates to the field of optical microscopy imaging technology, specifically to a wide field-of-view rotary confocal tomography imaging system. Background Technology

[0002] Optical microscopes have expanded human vision from the macroscopic to the microscopic realm, finding applications in numerous fields such as biomedicine and engineering. Among them, laser scanning confocal microscopy, with its optical tomography capabilities and high signal-to-noise ratio, has become the gold standard for observing subcellular microstructures. Traditional point-scanning confocal microscopy effectively blocks defocus signals by placing a pinhole in front of the detector, thereby accurately extracting focal plane information and significantly suppressing background noise. However, its point-by-point scanning mechanism severely limits imaging speed; simultaneously, the high-intensity illumination during long-term scanning can easily cause significant photobleaching and phototoxicity effects, which is detrimental to long-term observation of live samples.

[0003] To address the issues of improving imaging speed and reducing optical damage, rotating confocal microscopes were developed. They achieve parallel illumination and detection through a high-speed rotating array of microlenses and pinholes, significantly increasing frame rate and reducing optical damage, making them an important tool for live-cell imaging. However, limited by the design and manufacturing of imaging objectives, the field of view of traditional rotating confocal systems is typically limited to 1-2 millimeters, making it difficult to balance imaging speed and optical damage even for samples ranging from millimeters to centimeters in size.

[0004] Currently, the common challenges faced by rotating disk confocal microscopy imaging technology mainly lie in its limitations in the size of the imaging field of view, resolution, and imaging speed. In particular, existing technologies often struggle to achieve both large field of view and high-resolution imaging simultaneously: expanding the field of view usually leads to a decrease in spatial resolution, while pursuing high resolution necessitates shrinking the imaging field of view. This constraint severely limits the application of microscopy technology in high-resolution, detailed observation of large-sized samples. Utility Model Content

[0005] This invention addresses the problems in the prior art by disclosing a wide-field-of-view turntable confocal tomographic imaging system. This system combines an imaging objective with a traditional confocal structure, effectively overcoming the field-of-view limitations of traditional confocal microscopes. The second relay lens module effectively suppresses vignetting, a common phenomenon in multi-level relay imaging systems. Using a large-field imaging objective as the core imaging element, it supports high-resolution image acquisition within a centimeter-scale wide field of view, effectively solving the problem that expanding the field of view typically leads to a decrease in spatial resolution, while pursuing high resolution necessitates reducing the imaging field of view.

[0006] This utility model is achieved through the following technical solution: This invention first provides a wide-field-of-view turntable confocal tomography imaging system, comprising a housing and independently installed components including a laser source, a first relay lens module, a dichroic mirror, a pinhole turntable, a second relay lens module, a third relay lens module, and a detector. The first relay lens module and the dichroic mirror are arranged in the output light path of the laser source. The second relay lens module consists of two camera lenses forming the imaging objective, and the third relay lens module also consists of two camera lenses. Both the second and third relay lenses are combined in a configuration where the filter mounts face each other and the flanges face each other. The dichroic mirror has a pinhole turntable and a second relay lens module in its transmission light path; the second relay lens module is located above the sample, and the sample is located above the multi-degree-of-freedom adjustment component. A third relay lens module and a detector are provided in the reflected light path of the dichroic mirror; The second relay lens module and the third relay lens module constitute a conjugate optical module; The system is also equipped with a signal synchronization processing module for synchronizing the working timing of the pinhole disc, detector, and laser.

[0007] Preferably, the laser source matches the wide-field illumination to the current focal plane imaging area through the first relay lens module and the second relay lens module; the second relay lens module and the third relay lens module collect the returned signal light to the detector; the second relay lens module and the third relay lens module are used to realize triple conjugation and vignetting compensation; the triple conjugation is the conjugation between the pinhole turntable surface, the sample surface and the imaging surface.

[0008] Preferably, the second lens module, the sample, the third lens module, and the detector can be interchanged according to the transmission and reflection bands of the dichroic mirror.

[0009] Preferably, the housing is mounted on a height-adjustable bracket, and a mounting plate for mounting the detector is bolted to one side of the bracket.

[0010] Preferably, the laser source is emitted by a laser.

[0011] Preferably, the laser source emitted by the laser is transmitted via optical fiber.

[0012] Preferably, the laser is a single-wavelength laser.

[0013] Preferably, the center wavelength of the laser includes 405nm, 488nm, 561nm and 640nm.

[0014] Preferably, the first relay lens module includes a first imaging lens, a microlens array, and a second imaging lens, which are installed independently along the optical path in sequence; the first imaging lens serves as a collimating lens to collimate the micro-divergent beam emitted from the excitation source; the microlens array and the second imaging lens are a set of 4f relay lenses.

[0015] Preferably, the focal length of the first imaging lens is 200-300mm.

[0016] Preferably, the focal length of the first imaging lens is 220 mm.

[0017] Preferably, the microlens array and the second imaging lens have focal lengths of 4.5 mm and 50 mm, respectively, and a distance of 54.5 mm between them, and are connected to the housing using a cage plate structure.

[0018] Preferably, the dichroic mirror is disposed between the pinhole turntable and the first relay lens module; the pinhole turntable is disposed on the light output side of the first relay lens module.

[0019] Preferably, the dichroic mirror is a multi-band dichroic mirror.

[0020] Preferably, the dichroic mirror is a four-band dichroic mirror, which is fixedly mounted on the housing using a mirror frame, and its transmission wavelength is 397-413 nm / 482-492 nm / 555-567 nm / 630-645 nm.

[0021] Preferably, the pinhole turntable assembly includes a pinhole turntable and a connected rotary motor. The rotary motor is connected to a drive slider below, and the drive slider is installed above a linear rail. Bearing plates and motor flange plates are respectively installed at both ends of the linear rail. The drive slider is internally threaded with a lead screw. One end of the lead screw is rotatably connected to the bearing plate, and the other end is connected to the drive end of the motor. The rotary motor signal is connected to the microcontroller of the signal synchronization processing module, and the direction of motor movement is perpendicular to the optical axis of the imaging optical path.

[0022] Preferably, the motor is fixedly mounted on the housing.

[0023] Preferably, the pinhole turntable is configured with pinhole areas of different radii.

[0024] Preferably, the pinhole turntable is arranged in an Archimedean spiral pattern and has pinholes of different radii in different partitions. It can switch to the corresponding conjugate pinhole array for matching according to the imaging requirements of different samples, and can switch for imaging of samples of different thicknesses.

[0025] Preferably, the diameter of the light-transmitting pinholes on the pinhole turntable is between 5μm and 50μm, and the distance between adjacent pinholes is between 25μm and 500μm.

[0026] Preferably, the second relay lens module is disposed between the pinhole turntable and the sample; the second relay lens module includes a vignetting compensation lens, a first imaging objective lens, and a second imaging objective lens, which are installed independently in sequence from the dichroic mirror to the sample.

[0027] Preferably, the second relay lens module includes a vignetting compensation lens and a set of 4f first imaging objectives and second imaging objectives.

[0028] Preferably, the second relay lens module is composed of a first imaging objective with a focal length of 100 mm and a second imaging objective with a focal length of 50 mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate by double male rings.

[0029] Preferably, the vignetting compensation lens is a spherical lens with a focal length of 50 mm to 300 mm.

[0030] Preferably, the third relay lens module is disposed between the dichroic mirror and the detector; the third relay lens module includes a third imaging objective, a second filter, a fourth imaging objective, and a third filter, which are installed independently in sequence from the dichroic mirror to the detector.

[0031] Preferably, the third imaging objective and the second imaging objective are a pair of 4f systems with a spacing equal to the sum of their focal lengths. The focal lengths of both the third and fourth imaging objectives are 100 mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate via double male rings, and their ends are connected to the detector.

[0032] Preferably, the second and third filters are switchable filter modules.

[0033] Preferably, the second and third filters are filters with center wavelengths / bandwidths of 460 nm / 30 nm, 525 nm / 30 nm, 600 nm / 30 nm, and 670 nm / 20 nm, respectively.

[0034] Preferably, the detector is a two-dimensional area array camera.

[0035] The features and beneficial effects of this utility model are as follows: (1) This utility model combines imaging objective lens with traditional confocal structure, which effectively overcomes the field of view limitation of traditional confocal microscope. The second relay lens module used effectively suppresses the vignetting phenomenon common in multi-level relay imaging systems. The imaging objective lens with a large image field is used as the core imaging element, which supports high-resolution image acquisition in a centimeter-level large field of view. It effectively solves the problem that expanding the field of view usually leads to a decrease in spatial resolution, while pursuing high resolution requires shrinking the imaging field of view.

[0036] (2) The pinhole turntable of this invention is configured with a multi-sequence conjugate pinhole array according to the sample thickness, and effectively suppresses defocus background noise by means of the confocal pinhole structure, significantly improving the image signal-to-noise ratio. The turntable realizes rapid scanning and real-time optical tomography of the sample during high-speed rotation, greatly suppressing stray light interference.

[0037] (3) This utility model combines imaging objective lens and rotating disk confocal method to overcome the limitations of traditional rotating disk confocal in terms of field of view and imaging speed. At the same time, it effectively suppresses stray light and provides a high-throughput, cost-effective and easy-to-implement high-resolution imaging tool for life science and medical research. It has a large field of view, high resolution and fast tomography capability, and provides a complete solution for two-dimensional dynamic high-definition observation of large-size samples.

[0038] (4) The present invention optimizes the design of the relay optical path, which significantly improves the uniformity of the illumination distribution in the field of view, thereby improving the signal consistency and image availability in the edge area while ensuring the overall imaging quality. Attached Figure Description

[0039] To more clearly illustrate the technical solutions in the embodiments of this utility model, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this utility model. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0040] Figure 1 This is a schematic diagram of the principle of the wide field-of-view turntable confocal tomography imaging system described in this embodiment of the present invention; Figure 2 This is a diagram showing the positional relationship of the optical modules described in this embodiment of the invention. Figure 3 This is a schematic diagram of the installation of the optical module within the housing according to an embodiment of the present invention; Figure 4 This is a front view of the system described in an embodiment of the present utility model; Figure 5 This is a side view of the system described in an embodiment of the present utility model; Figure 6 This is a schematic diagram of the pinhole turntable assembly described in an embodiment of the present utility model; Figure 7 This is a schematic diagram of the pinhole turntable described in an embodiment of the present utility model; Figure 8 This is a schematic diagram of the driving unit described in an embodiment of the present utility model; Figure 9 This is a low-frequency timing control diagram for the signal synchronization processing module described in this embodiment of the utility model; Figure 10 This is a high-frequency timing control diagram for the signal synchronization processing module described in this embodiment of the present invention.

[0041] Explanation of reference numerals in the attached figures: 1-Housing; 2-Placement stage; 3-Swivel mechanism; 4-Vertical lifting platform; 5-Horizontal translation stage; 6-Support rod; 7-Optical plate; 8-Pressure plate; 9-Support base; 10-Support component; 11-Laser; 12-Mounting plate; 13-Fiber optic cable; 14-First relay lens module; 15-Dichroic mirror; 16-Pinhole turntable; 17-Second relay lens module; 18-Sample; 19-Third relay lens module; 20-Detector; 21-Pressure plate; 22-Rotary motor; 23-Connector; 24-Motor; 25-Motor flange; 26-Linear rail; 27-Bearing plate; 28-Lead screw; 29-Drive slider. Detailed Implementation

[0042] To facilitate understanding of this utility model, a more comprehensive description of this utility model will be provided below, along with embodiments of this utility model, but this does not limit the scope of this utility model.

[0043] In the description of this utility model, it should be understood that the terms "center," "longitudinal," "lateral," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," and "outer," etc., indicating orientation or positional relationships based on the orientation or positional relationships shown in the accompanying drawings, are only for the convenience of describing this utility model and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this utility model. Furthermore, the terms "first," "second," etc., are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of indicated technical features. Thus, features defined with "first," "second," etc., may explicitly or implicitly include one or more of that feature. In the description of this utility model, unless otherwise stated, "a plurality of" means two or more.

[0044] In the description of this utility model, it should be noted that, unless otherwise explicitly specified and limited, the terms "installation," "connection," and "joining" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal connection of two components. Those skilled in the art can understand the specific meaning of the above terms in this utility model based on the specific circumstances.

[0045] A wide-field-of-view rotary confocal tomography imaging system includes mechanical modules (such as housing, motor, and five-degree-of-freedom adjustment components) for auxiliary optical modules and sample mounting. Figures 1 to 10 As shown, it also includes a housing 1 and an optical module installed inside it. The optical module includes a laser source, a first relay lens module 14, a dichroic mirror 15, a pinhole turntable 16, a second relay lens module 17, a third relay lens module 19, and a detector 20, which are installed independently (bolted or bolted together by connectors). The laser source has a first relay lens module 14 and a dichroic mirror 15 installed in its output light path. The dichroic mirror 15 has a pinhole turntable 16 and a second relay lens module 17 on its transmission light path; the second relay lens module 17 is located above the sample 18, and a five-degree-of-freedom adjustment component for adjusting multiple dimensional directions is installed below the sample 18. The dichroic mirror 15 is provided with a third relay lens module 19 and a detector 20 in the reflected light path; The second relay lens module 15 and the third relay lens module 19 constitute a conjugate optical module; The second relay lens module 15 is composed of two camera lenses forming an imaging objective lens, and the third relay lens module is formed by combining two camera lenses; both the second and third relay lenses are combined in a manner where "filter mounts face each other and flange mounts face each other"; The system also includes a signal synchronization processing module for synchronizing the operating timing of the pinhole disc, detector, and laser.

[0046] The pinhole turntable assembly includes a pressure plate 21, a pinhole turntable 16, a rotary motor 22, and a connector 23. The drive unit includes a motor 24, a motor flange 25, a linear guide rail 26, a bearing plate 27, a lead screw 28, and a drive slider 29. Bolts pass sequentially through the mounting holes of the pinhole turntable 16 and the pressure plate 21, and are then fixedly connected to the movable end of the rotary motor 22. The fixed end of the rotary motor 22 is connected to the drive slider 29 via the connector 23. The drive slider 29 is mounted above the linear guide rail 26. The bearing plate 27 and the motor flange 25 are respectively mounted at both ends of the linear guide rail 26. The drive slider 29 is internally threadedly connected to the lead screw 28. One end of the lead screw 28 is rotatably connected to the bearing plate 27, and the other end passes through the motor flange 15 and is connected to the drive end of the motor 24. Both the rotary motor 22 and the motor 24 are signal-connected to the microcontroller of the signal synchronization processing module. The motor 24 is fixedly mounted perpendicular to the optical axis of the imaging optical path. Controlled by the signal synchronization processing module, it drives the pinhole turntable to move radially, thereby precisely switching the pinholes of different sizes into the optical path's active area to adjust the imaging field of view or tomographic capability. The drive unit is fixedly mounted on the housing 1. The rotary motor 22 is a three-phase high-speed brushless DC motor that receives control commands from the signal synchronization processing module to adjust the turntable speed in real time.

[0047] In one embodiment, a multi-degree-of-freedom adjustment assembly is used to achieve three-dimensional linear translation and biaxial deflection around a fixed point in space. It includes a tilting device 3, a vertical lifting platform 4, and a horizontal translation platform 5, bolted together from top to bottom. The tilting device 3 includes an X-axis tilting device and a Y-axis tilting device vertically connected below it; the horizontal translation platform 5 includes an X-axis translation platform and a Y-axis translation platform vertically connected below it. The tilting device 3, vertical lifting platform 4, and horizontal translation platform 5 all have a sliding block structure, and a knob for fixing the slider is provided on one side of the sliding block. To facilitate the placement of the sample 18, a sample placement platform 2 is threaded onto the top of the tilting device 3. The horizontal translation platform 5 is connected to an optical plate 7 via a support rod 6. The optical plate 7 is also used to place a laser 11. The vertical lifting platform 4 is model LZ60. The tilting device 3 is model GFG60. The horizontal translation platform is model E-EIL36-90.

[0048] In one embodiment, the multi-degree-of-freedom adjustment component is a five-dimensional adjustment frame as described in the prior art.

[0049] Preferably, the support rod 6 is sleeved with the support base 8 below, and is connected to the optical plate 7 through the support bracket 9.

[0050] Preferably, the housing 1 is mounted on a bracket, the bracket height is adjustable so that the installation height of the housing is adjustable, and it is fixed by a support member 10. One side of the profile is bolted to a mounting plate 12, which is used to install the detector 20. This makes the structure of this application more compact and allows for vertical installation, which is conducive to maximizing space utilization and saving floor space.

[0051] In some embodiments, the laser source is emitted by laser 11; Preferably, the laser source emitted by the laser 11 is transmitted through the optical fiber 13.

[0052] In some embodiments, the laser 11 is a single-wavelength laser with narrow linewidth and high stability, used to excite sample fluorescence; it can illuminate the imaging area corresponding to the focal position during the scanning cycle of the pinhole turntable.

[0053] In some embodiments, the center wavelength of the laser 11 includes, but is not limited to, 405nm, 488nm, 561nm, and 640nm.

[0054] In some embodiments, the first relay lens module 14 includes a first imaging lens 141, a microlens array 142, and a second imaging lens 143, which are independently mounted sequentially along the optical path (from the laser source to the dichroic mirror 15). The first imaging lens 141 serves as a collimating lens for collimating the micro-divergent beam emitted from the excitation source. The microlens array 142 and the second imaging lens 143 form a set of 4f relay lenses for homogenizing and expanding the beam to provide uniform illumination and match the aperture requirements of subsequent optical paths. In some embodiments, the first imaging lens 141 has a focal length of 200-300 mm, preferably 220 mm, to collimate the micro-divergent beam emitted from the excitation source, ensuring parallel beam transmission between groups; the microlens array 142 and the second imaging lens 143 have focal lengths of 4.5 mm and 50 mm, respectively, with a spacing of 54.5 mm, and are fixedly mounted on the housing 1 using a cage plate structure, for speckle removal and beam expansion, to provide uniform illumination and match the aperture requirements of the subsequent optical path, thereby achieving uniform filling of the back focal plane of the objective lens and large-area illumination coverage of the sample 18.

[0055] Dichroic mirror 15 is disposed between pinhole turntable 16 and first relay lens module 14; pinhole turntable 16 is disposed on the light output side of first relay lens module 14.

[0056] In some embodiments, the dichroic mirror adopts a multi-band design, which can be compatible with a variety of fluorescent dyes at the same time.

[0057] In some embodiments, the pinhole turntable, the second lens module, the sample, the third lens module, and the detector can be interchanged according to the transmission and reflection bands of the dichroic mirror.

[0058] In some embodiments, the dichroic mirror 15 is a four-band dichroic mirror, which is fixedly mounted on the housing 1 with a mirror frame. Its transmission wavelength is 397-413 nm / 482-492 nm / 555-567 nm / 630-645 nm. It is used for the excitation and detection of different fluorescent dyes in the sample 18, thereby realizing the separation of the optical path for excitation illumination and signal collection.

[0059] In some embodiments, the pinhole turntable 16 uses pinholes arranged in an Archimedean spiral pattern and configured with different radii in different partitions. It can switch to the corresponding conjugate pinhole array for matching according to the imaging requirements of different samples, and can switch for imaging samples of different thicknesses. It is used to switch pinholes of different diameters to adjust the depth of field and tomographic capability of the system, so as to selectively collect light signals from different depths near the focal plane, thereby adapting to the imaging requirements of thin, medium and thick samples.

[0060] In some embodiments, pinhole areas of different radii are configured on the pinhole turntable 16.

[0061] In some embodiments, the pinhole turntable 16 can be fabricated using a chrome plate masking process.

[0062] In some embodiments, the diameter of the light-transmitting pinholes of the pinhole turntable 16 is between 5 micrometers and 50 micrometers, and the distance between adjacent pinholes is between 25 micrometers and 500 micrometers.

[0063] In some embodiments, a second relay lens module 17 is disposed between the pinhole turntable 16 and the sample 18; the second relay lens module 17 includes a vignetting compensation lens 171, a first imaging objective lens 172 and a second imaging objective lens 173, which are installed independently in sequence from the dichroic mirror 15 to the sample 18, to achieve uniform, large field of view illumination and imaging.

[0064] In some embodiments, the second relay lens module 17 is an optional vignetting compensation lens 171 and a set of 4f first imaging objectives 172 and second imaging objectives 173. The first imaging objectives 172 and second imaging objectives 173 serve as the core imaging components of the module, and their large aperture and low distortion optical design ensures high resolution and wide illumination throughout the entire field of view.

[0065] In some embodiments, the second relay lens module 17 consists of a first imaging objective lens 172 with a focal length of 100 mm and a second imaging objective lens 173 with a focal length of 50 mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate by double male rings; together they constitute the imaging objective lens to realize wide field of view illumination and imaging of the sample surface.

[0066] The function of the vignetting compensation lens 171 is to eliminate vignetting caused by cascaded relay lenses. When multiple relay lenses are used in cascade, changes in the angle of light may cause darkening or blurring of image edges. The role of the vignetting compensation lens is to eliminate this vignetting caused by cascaded relay lenses, ensuring consistency and sharpness of image quality.

[0067] In some embodiments, the vignetting compensation lens 171 may be implemented by placing a spherical lens with a focal length of 50 mm to 300 mm near the conjugate relay plane of the image plane.

[0068] In some embodiments, the third relay lens module 19 includes a pair of camera lenses with a focal length of 100 mm as a relay lens. The filter mounts of the two objectives are connected and fixed to the optical cage plate by a double male ring, which is used to extend the physical working distance of the imaging system and provide the image-side telecentric optical path for the detector.

[0069] In some embodiments, a third relay lens module 19 is disposed between the dichroic mirror 15 and the detector 20; the third relay lens module 19 includes a third imaging objective 191, a second filter 192, a fourth imaging objective 193, and a third filter 194, which are installed independently in sequence from the dichroic mirror 15 to the detector 20; the third and fourth imaging objectives are used to extend the physical working distance of the optical path of the imaging system; the second and third filters are switchable filter modules to adapt to sample imaging of different fluorescence bands, and are used to suppress stray light reflected and scattered from all directions, thereby improving the signal-to-noise ratio of the image; In some embodiments, the third imaging objective 191 and the second imaging objective 173 in the second intermediate lens module 17 form a 4f system, with a spacing equal to the sum of their focal lengths. Both the third and fourth imaging objectives have a focal length of 100mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate via double male rings, and their ends are connected to the detector 20 via a camera interface. This module effectively extends the working distance of the imaging optical path, providing the detector with ample imaging space and a uniform light field distribution.

[0070] In some embodiments, the second filter 192 and the third filter 193 are switchable and selected according to the corresponding detection fluorescence wavelength to filter out stray light generated by reflection from the sample surface, etc. Example configurations include filters with center wavelengths / bandwidths of 460 nm / 30 nm, 525 nm / 30 nm, 600 nm / 30 nm, and 670 nm / 20 nm, respectively.

[0071] In some embodiments, the detector 20 is a two-dimensional area array camera used to read the excitation fluorescence signal on the sample and store the collected information.

[0072] In some embodiments, the laser source 10 matches the wide-field illumination to the current focal plane imaging region through the first relay lens module 14 and the second relay lens module 17; the second relay lens module 17 and the third relay lens module 19 collect the returned signal light to the detector 20; the second relay lens module 17 and the third relay lens module 19 are used to realize triple conjugation and vignetting compensation; the triple conjugation is the conjugation between the pinhole turntable surface, the sample surface and the imaging surface.

[0073] In some embodiments, depending on the excitation and emission bands of different fluorescent samples, the laser light source, dichroic mirror, and filter can be selectively replaced to adapt to sample imaging in different fluorescence bands.

[0074] The signal synchronization processing module is used to synchronize the working timing of the pinhole turntable 16, detector 20, and laser 13, and to synchronously control the acquisition timing of the detection module according to the phase signal. The signal synchronization processing module includes a microcontroller for synchronously sampling the trigger signal and a photoelectric sensor for detecting the phase of the pinhole turntable 16. The photoelectric sensor is installed inside the housing 1 and located on one side of the pinhole turntable 16. The photoelectric sensor is an H206 slot-type optocoupler. The microcontroller is an STM8S103F3P6 microcontroller. The microcontroller is connected to the photoelectric sensor, rotary motor 22, motor 24, detector 20, and an external host computer. The microcontroller synchronously samples the signal acquired by the photoelectric sensor to calculate the real-time rotation speed and feeds it back to the host computer, thereby setting the exposure time of detector 20 to an integer multiple of the turntable cycle to ensure acquisition synchronization accuracy. Simultaneously, the microcontroller generates a fast high-level pulse (duration 5ms) through its GPIO port as a trigger signal to control detector 20 to capture images and perform real-time preview. This synchronously triggers the detector to perform exposure, thereby achieving high-precision synchronous operation of the entire system. The microcontroller, photoelectric sensor, rotary motor 22, motor 24, and detector 20 all utilize existing chips and motors, and their specific connection methods and communication methods are also existing technologies. Furthermore, the inventive point of this paper primarily lies in the improvement of the mechanical structure, without involving improvements to circuitry or communication technology; therefore, these will not be elaborated upon further. The signal synchronization processing module also includes a power supply unit, which can be a commonly used power supply unit in the prior art, primarily supplying power to the rotary motor 22, motor 24, and microcontroller. The rotary motor 22 is a JS30 model, and the motor 24 is a 50KTYZ model. The detector 20 is a high-sensitivity sCMOS camera capable of acquiring signal light in real time over a large field of view, suitable for tomographic imaging applications that are sensitive to noise and require high image quality.

[0075] Preferably, the signal detected by the detector can be a reflected signal, a scattered signal, or a fluorescence signal from the focal point. The signal light returns through the imaging system and passes through the pinhole again. At this point, the pinhole blocks most of the stray light from other areas, allowing only the signal from the laser focusing area to pass through efficiently. For example... Figure 3 and Figure 4 As shown, by synchronizing the laser illumination trigger signal with the signals of the turntable and detector 20, the illumination light can be precisely covered within the exposure time window of the detector 20. Specifically, for high-speed shooting modes (e.g., 100Hz), the laser source can use continuous illumination; while for shooting dynamic samples, a low-frequency signal can be used to trigger the turntable and camera signals (e.g., 50-70 Hz), and the laser source switches to on-demand illumination (i.e., illumination only during exposure). This on-demand illumination method can reduce the total light dose, effectively ensuring low phototoxicity, thereby achieving low-damage long-term shooting of live samples.

[0076] In some embodiments, the turntable generates at least one trigger signal per revolution, the trigger frequency being proportional to the turntable rotation speed; the system can be configured to trigger the turntable multiple times to activate the camera to perform single or multiple exposures, or to make the camera exposure time proportional to the set number of trigger cycles, thereby achieving flexible imaging timing control.

[0077] The housing 1 is made of black anodized aluminum with a thickness of millimeters, and the base plate is an optical breadboard with dimensions of 300 mm × 420 mm.

[0078] This invention implements a wide-field-of-view rotating disk confocal microscopy tomography method. Building upon traditional rotating disk confocal microscopy, it utilizes a vignetting compensation lens and a large-image-field photographic lens as imaging objectives to create a broad and uniform light field coverage on the sample. Simultaneously, a signal synchronization processing module coordinates the timing of the pinhole rotating disk and the detection module, enabling imaging of samples of varying thicknesses using a pre-set multi-sequence conjugate pinhole array. This approach significantly expands the field of view while maintaining high resolution and optical tomographic capabilities in confocal imaging, achieving high-speed, large-area, high-resolution imaging.

[0079] It should be noted that the above description is only a preferred embodiment of the present utility model and is not intended to limit the present utility model. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present utility model should be included within the protection scope of the present utility model.

Claims

1. A wide-field-of-view rotating disk confocal tomography imaging system, characterized in that: It includes a housing and its internal components, each independently mounted, such as a laser source, a first relay lens module, a dichroic mirror, a pinhole turntable, a second relay lens module, a third relay lens module, and a detector; wherein, the laser source's output light path is provided with the first relay lens module and the dichroic mirror; The second relay lens module consists of two camera lenses forming the imaging objective, and the third relay lens module consists of two camera lenses; both the second and third relay lenses are combined in a manner where the filter mounts face each other and the flange mounts face each other. The dichroic mirror has a pinhole turntable and a second relay lens module in its transmission light path; the second relay lens module is located above the sample, and the sample is located above the multi-degree-of-freedom adjustment component. A third relay lens module and a detector are provided in the reflected light path of the dichroic mirror; The second relay lens module and the third relay lens module constitute a conjugate optical module; The system also includes a signal synchronization processing module for synchronizing the operating timing of the pinhole disc, detector, and laser.

2. The wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The laser source matches the wide-field illumination to the current focal plane imaging area through the first relay lens module and the second relay lens module; the second relay lens module and the third relay lens module collect the returned signal light to the detector; The second and third relay lens modules are used to achieve triple conjugation and vignetting compensation; the triple conjugation is the conjugation between the pinhole turntable surface, the sample surface, and the imaging surface. Preferably, the second lens module, the sample, the third lens module, and the detector can be interchanged according to the transmission and reflection bands of the dichroic mirror.

3. The wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The housing is mounted on a height-adjustable bracket, with a mounting plate for mounting the detector bolted to one side of the bracket.

4. The wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The laser source is emitted by a laser; Preferably, the laser source emitted by the laser is transmitted via optical fiber; Preferably, the laser is a single-wavelength laser; Preferably, the center wavelength of the laser includes 405nm, 488nm, 561nm and 640nm.

5. The wide field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The first relay lens module includes a first imaging lens, a microlens array, and a second imaging lens, which are installed independently along the optical path in sequence. The first imaging lens serves as a collimating lens to collimate the micro-divergent beam emitted from the excitation source. The microlens array and the second imaging lens form a set of 4f relay lenses. Preferably, the focal length of the first imaging lens is 200-300mm. Preferably, the focal length of the first imaging lens is 220 mm; Preferably, the microlens array and the second imaging lens have focal lengths of 4.5 mm and 50 mm, respectively, and a distance of 54.5 mm between them, and are connected to the housing using a cage plate structure.

6. The wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: A dichroic mirror is disposed between the pinhole turntable and the first relay lens module; the pinhole turntable is disposed on the light output side of the first relay lens module. Preferably, the dichroic mirror is a multi-band dichroic mirror; Preferably, the dichroic mirror is a four-band dichroic mirror, which is fixedly mounted on the housing using a mirror frame, and its transmission wavelength is 397-413 nm / 482-492 nm / 555-567 nm / 630-645 nm.

7. The wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The pinhole turntable assembly includes a pinhole turntable and a connected rotary motor. The rotary motor is connected to a drive slider below. The drive slider is mounted above a linear rail. Bearing plates and motor flange plates are respectively installed at both ends of the linear rail. The drive slider is internally threaded with a lead screw. One end of the lead screw is rotatably connected to the bearing plate, and the other end is connected to the drive end of the motor. The rotary motor signal is connected to the microcontroller of the signal synchronization processing module, and the direction of the motor's movement is perpendicular to the optical axis of the imaging optical path. Preferably, the motor is fixedly mounted on the housing; Preferably, the pinhole turntable is configured with pinhole areas of different radii; Preferably, the pinhole turntable is arranged in an Archimedean spiral pattern and has pinholes of different radii in different partitions. It can switch to the corresponding conjugate pinhole array for matching according to the imaging requirements of different samples, and can switch for imaging of samples of different thicknesses. Preferably, the diameter of the light-transmitting pinholes on the pinhole turntable is between 5 micrometers and 50 micrometers, and the distance between adjacent pinholes is between 25 micrometers and 500 micrometers.

8. The wide field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The second relay lens module is disposed between the pinhole turntable and the sample; the second relay lens module includes a vignetting compensation lens, a first imaging objective lens, and a second imaging objective lens, which are installed independently in sequence from the dichroic mirror to the sample. Preferably, the second relay lens module includes a vignetting compensation lens and a set of 4f first imaging objectives and second imaging objectives; Preferably, the second relay lens module is composed of a first imaging objective with a focal length of 100 mm and a second imaging objective with a focal length of 50 mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate by double male rings. Preferably, the vignetting compensation lens is a spherical lens with a focal length of 50 mm to 300 mm.

9. A wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The third relay lens module is disposed between the dichroic mirror and the detector; the third relay lens module includes a third imaging objective, a second filter, a fourth imaging objective, and a third filter, which are installed independently in sequence from the dichroic mirror to the detector; Preferably, the third imaging objective and the second imaging objective are a pair of 4f systems with a spacing equal to the sum of their focal lengths. The focal lengths of both the third and fourth imaging objectives are 100 mm. The filter mounts of the two objectives are connected and fixed to the optical cage plate via double male rings, and their ends are connected to the detector. Preferably, the second and third filters are switchable filter modules; Preferably, the second and third filters are filters with center wavelengths / bandwidths of 460 nm / 30 nm, 525 nm / 30 nm, 600 nm / 30 nm, and 670 nm / 20 nm, respectively.

10. A wide-field-of-view rotating disk confocal tomography imaging system according to claim 1, characterized in that: The detector is a two-dimensional area array camera.

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