Confocal microscope illumination path for spinning disk parallel scanning

Abstract

The present disclosure describes a rotary disk parallel scanning confocal microscope illumination light path, which comprises a sleeve assembly with a light source and a field diaphragm, a lens barrel assembly with a condenser lens, a light splitting chamber with a rotary disk, a tube lens and a microscope objective, the light source is configured to form a first parallel light beam entering the lens barrel assembly via the field diaphragm; the light source is imaged at the focal plane of the condenser lens; the rotary disk is configured to receive a second parallel light beam emitted from the tube lens, the second parallel light beam is configured to form a preset image on the rotary disk, the rotary disk is configured to receive the first parallel light beam emitted from the lens barrel assembly into the light splitting chamber, the first parallel light beam is configured to form a light spot on the rotary disk covering the aperture surface of the rotary disk; the microscope objective is configured to be installed at the position of the image of the light source formed by the light beam emitted from the tube lens, which meets the preset requirements. According to the present disclosure, a rotary disk parallel scanning confocal microscope illumination light path which is easy to operate and can be easily and accurately adjusted can be provided.

Description

[0001] This application is a divisional application of the patent application filed on August 26, 2022, with application number 2022110299739, entitled "Method for Adjusting Illumination Optical Path of Microscope". Technical Field

[0002] This disclosure generally relates to an intelligent manufacturing equipment industry, specifically to an illumination optical path for a rotary parallel scanning confocal microscope. Background Technology

[0003] Currently, optical microscopy is widely used in various fields of scientific and technological research. However, ordinary optical microscopy cannot achieve three-dimensional imaging of objects with a certain thickness. With the continuous development of microscopy technology in recent years, confocal microscopy has become one of the important technologies in the field of optical microscopy. It has the characteristics of high precision, high resolution, non-contact, and unique axial tomographic scanning imaging, which can easily realize three-dimensional image reconstruction and has been widely used in micro-nano detection, precision measurement, and life science research. Traditional confocal microscopy detection technology is based on the principle of single-point mechanical scanning of the light source, the illuminated object point, and the detector. Therefore, the scanning speed is relatively slow, the mechanical control system is complex, and the vibration caused by scanning limits the measurement accuracy, making it difficult to achieve real-time and rapid three-dimensional measurement. To solve the shortcomings of slow imaging speed and small field of view of confocal microscopes, parallel scanning confocal microscopy technology has emerged. Parallel scanning confocal microscopy technology improves the measurement speed of the original single-point confocal measurement, and parallel scanning confocal microscopy detection technology based on Nipkow turntable has the advantages of simple structure, easy implementation, low cost, and high image quality.

[0004] In the Nipkow rotary parallel scanning confocal microscope, the illumination optical path and the detection optical path need to share the small hole on the Nipkow rotary disk. At this time, the reasonable setting and accurate adjustment of the illumination optical path are particularly important. The illumination optical path is one of the essential optical paths in microscopy. Ensuring that the illumination optical path has good and uniform illumination conditions is a necessary condition for achieving high-precision and high-resolution measurements.

[0005] Patent document [CN110764271A] discloses a method for adjusting the positional accuracy between lenses in a conjugate optical system. It utilizes a high-precision air-bearing pivot to replace the focusing axis of an internal focusing telescope, enabling the lens optical axis adjustment accuracy to reach the micrometer level. However, the solution disclosed in this patent document is costly and complex to operate, making it unsuitable for assembling and adjusting the illumination optical path of a parallel scanning confocal microscope. Summary of the Invention

[0006] This disclosure was made in view of the aforementioned state of the prior art, and its purpose is to provide a method for easily operating and accurately adjusting the illumination optical path of a rotary parallel scanning confocal microscope. By utilizing a reasonably designed illumination optical path structure and accurately adjusting the position of the observation window, the illumination optical path can be finely calibrated step by step in sections, thereby improving the illumination accuracy of the light source in the microscope imaging system and perfecting the imaging quality of the microscopic detection optical path.

[0007] Therefore, this disclosure provides a method for assembling and adjusting a microscope illumination optical path. The microscope illumination optical path includes a sleeve assembly with a light source and a field aperture, a microscope tube assembly with a condenser and an aperture stop, a beam splitter and a rotating disk with a small aperture, a microscope tube, and a microscope objective. The method for assembling and adjusting the illumination optical path includes: setting the light source to form a first parallel beam entering the microscope tube assembly via the field aperture; setting the aperture stop at the focal plane of the condenser; adjusting the orientation of the light source so that the light source forms a centrally symmetrical image in the aperture stop; setting the rotating disk and the beam splitter in the beam splitter; forming... A second parallel beam is directed from the tube lens into the turntable of the beam splitter. The relative position of the tube lens and the beam splitter is adjusted so that the second parallel beam forms a preset image on the turntable. A first parallel beam is directed through the lens assembly into the beam splitter and forms a light spot on the turntable. The relative position of the field stop and the condenser lens is adjusted so that the light spot covers the aperture surface. Images of the light source formed by the beam emitted from the tube lens are received at different positions. Based on the position of the image of the light source that meets the preset requirements, the installation position of the microscope objective is determined and the microscope objective is installed.

[0008] The assembly method disclosed herein is based on Köhler illumination requirements, adhering to the illumination condition of "pupil to window, window to pupil." The effect of this illumination condition is to image the light source on the image-side back focal plane of the microscope objective, and to achieve a conjugate relationship between the field stop and the object-side focal plane of the microscope objective. As a result, the surface of the illuminated object sample is smoothly and uniformly illuminated without any shadows.

[0009] The adjustment method disclosed herein first collimates the light beam of the light source, focusing the edge rays to emit them as parallel light, thereby homogenizing the light energy distribution within the cross-section perpendicular to the optical axis. Then, the light passes through the field stop and the focusing lens, forming an image of the light source at the aperture stop. Adjusting the orientation of the light source ensures that it forms a centrally symmetrical image within the aperture stop, enabling the optical axis of the parallel beam to pass through the center of the aperture stop. Furthermore, adjusting the orientation of the light source ensures no loss of illumination energy, thus reducing aberrations caused by the optical axis not passing through the center of the aperture stop. Furthermore, by adjusting the relative positions of the field stop, the aperture stop, the turntable, and the tube lens in segments and sections, the image of the light source formed by the light beam emitted from the tube lens is received. The imaging position of the image of the light source that meets the preset requirements is the installation position of the image-side back focal plane of the microscope objective. In this case, the effect of imaging the light source on the image-side back focal plane of the microscope objective can be achieved. At the same time, by making the field stop image cover the small aperture surface of the turntable, the effect of the field stop and the object-side focal plane of the microscope objective being conjugate can be achieved.

[0010] Furthermore, in the assembly method of this embodiment, optionally, the field stop has a non-circular through-hole, and the field stop is a square aperture. This makes it easy to observe the centrally symmetrical image of the light source formed in the aperture stop, and the external image of the field stop on the small hole surface of the turntable.

[0011] In addition, in the assembly and adjustment method of this embodiment, optionally, the sleeve assembly includes a collimating lens, and the relative position of the light source and the collimating lens is adjusted to form a first parallel beam entering the lens assembly through the collimating lens. In this case, the collimating lens can effectively correct edge aberrations of the field of view, reduce spherical aberration, and improve light energy utilization.

[0012] Furthermore, in the assembly method described in this embodiment, optionally, an observation window is disposed at the aperture stop to receive the image of the light source formed by the light source at the aperture stop. Since it is difficult to obtain an image of the light source when the light beam passes through the through-hole of the aperture stop, by using an observation window disposed at the aperture stop, it is possible to obtain the light beam passing through the through-hole of the aperture stop to form an image of the light source, thereby enabling the determination of the relative position of the aperture stop and the condenser lens.

[0013] Furthermore, in the mounting method of this embodiment, optionally, an image of the light source formed by the light beam emitted from the tube lens is received using an observation window, and the observation window is moved along the optical axis until an image of the light source that meets preset requirements appears on the observation window. This allows the position of a clear light source image to be determined, thereby enabling the determination of the mounting position of the image-side back focal plane of the microscope objective. Consequently, the mounting position of the microscope objective can be determined based on the numerical relationship between the image-side focal plane and the parfocal plane of the microscope objective.

[0014] Alternatively, in the assembly method described in this embodiment, the observation window may be a semi-transparent object, specifically a semi-transparent frosted glass. In this case, the image of the light source formed by the aperture stop and the image of the light source formed by the light beam emitted from the tube lens are imaged onto the semi-transparent object, thereby facilitating visual observation of the corresponding imaging.

[0015] Furthermore, in the assembly and adjustment method of this embodiment, optionally, after adjusting the relative positions of the light source and the collimating lens, the positional relationship between the light source and the collimating lens is fixed; after adjusting the relative positions of the field stop and the condenser lens, the positional relationship between the field stop and the condenser lens is fixed; after adjusting the relative positions of the tube lens and the beam splitter, the positional relationship between the tube lens and the beam splitter is fixed. In this case, zonal assembly and adjustment can be achieved, ensuring that other assembly and adjustment steps do not affect the relative positional relationship between the already assembled and adjusted modules during the assembly and adjustment process.

[0016] Additionally, in the assembly and adjustment method of this embodiment, optionally, the sleeve assembly includes an outer rotating cylinder, and the side wall of the outer rotating cylinder is provided with an adjustment device for adjusting the position of the light source in the vertical plane of the optical axis. The adjustment device includes a set screw. Thus, by using this adjustment device, the orientation of the light source can be adjusted so that the light source forms a centrally symmetrical image in the aperture stop.

[0017] Additionally, in the assembly and adjustment method described in this embodiment, optionally, the second parallel beam is obtained through a collimator, and the preset image is a collimator reticle image. Since the reticle image is imaged at infinity by two parallel beams, the collimator can simulate the reflected beam from the sample being tested entering the turntable through the microscope objective, thereby enabling more accurate adjustment of the relative position of the collimator and the spectrometer.

[0018] Additionally, in the application device according to this embodiment, optionally, the sleeve assembly and the lens barrel assembly are connected by threads, and the tube lens and the beam splitter are connected by threads. Thus, the threaded connection allows the sleeve assembly to rotate along the threads, moving it along the optical axis, adjusting the relative position between the sleeve assembly and the lens barrel assembly, thereby adjusting the relative position between the field stop and the condenser lens; the threaded connection also allows the tube lens and the beam splitter to rotate along the threads, moving the tube lens along the optical axis, adjusting the relative position between the tube lens and the beam splitter, thereby adjusting the relative position between the tube lens and the turntable.

[0019] According to this disclosure, a method is provided that is easy to operate and can accurately adjust the illumination optical path of a rotary parallel scanning confocal microscope. By setting a reasonable illumination optical path structure and accurately adjusting the position of the observation window, and gradually completing the illumination optical path adjustment according to the partition, the illumination accuracy of the light source in the microscope imaging system can be improved, and the imaging quality of the microscopic detection optical path can be perfected. Attached Figure Description

[0020] Figure 1 This is a schematic diagram showing the optical path of the microscope involved in this embodiment example.

[0021] Figure 2 This is a schematic diagram showing the illumination optical path of the microscope involved in this embodiment example.

[0022] Figure 3 This is a schematic diagram showing the structure of the illumination optical path of the microscope involved in this embodiment example.

[0023] Figure 4A This is a flowchart illustrating a first embodiment of the assembly and adjustment method described in this example.

[0024] Figure 4B This is a flowchart illustrating a second embodiment of the assembly and adjustment method described in this example.

[0025] Figure 5 This is a schematic diagram illustrating a scenario for acquiring a first parallel beam as described in this embodiment example.

[0026] Figure 6 This is a schematic diagram illustrating a scene involving adjusting the posture of a light source as described in this embodiment example.

[0027] Figure 7 This is a schematic diagram showing the relative positions of the adjustment tube and the beam splitter in this embodiment example.

[0028] Figure 8 This is a schematic diagram illustrating a scenario involving adjusting the position of a microscope objective as described in this embodiment example. Detailed Implementation

[0029] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0030] It should be noted that the terms "first," "second," "third," and "fourth," etc., in the specification, claims, and accompanying drawings of this invention are used to distinguish different objects, not to describe a specific order. Furthermore, the terms "comprising" and "having," and any variations thereof, are intended to cover non-exclusive inclusion. For example, a process, method, system, product, or device that includes a series of steps or units is not limited to the listed steps or units, but may optionally include steps or units not listed, or may optionally include other steps or units inherent to these processes, methods, products, or devices. In the following description, the same reference numerals are used for the same parts, and repeated descriptions are omitted. Additionally, the accompanying drawings are merely schematic diagrams, and the scale of the dimensions of the parts or the shape of the parts may differ from the actual figures.

[0031] The embodiments of this disclosure relate to a method for assembling and adjusting the illumination optical path of a rotary parallel scanning confocal microscope. Using the assembly and adjustment method disclosed herein, the precise assembly and adjustment of the illumination optical path can be completed step-by-step, section by section, without the use of special instruments or equipment. The overall operation process is simple and easy to troubleshoot. The "method for assembling and adjusting the illumination optical path of a rotary parallel scanning confocal microscope" can also be referred to as an "assembly and adjustment method".

[0032] The assembly and adjustment method described in this embodiment will now be explained in detail with reference to the accompanying drawings.

[0033] Figure 1 This is a schematic diagram showing the optical path of the microscope involved in this embodiment example. Figure 2 This is a schematic diagram showing the illumination optical path of the microscope according to the example of this embodiment. Figure 3 This is a schematic diagram showing the structure of the illumination optical path of the microscope involved in this embodiment example.

[0034] In some examples, the microscope involved in this disclosure may be a parallel scanning confocal microscope. In some examples, the microscope involved in this disclosure may be a Nipkow rotary parallel scanning confocal microscope.

[0035] See in some examples Figure 1 The optical path of a microscope can include an illumination optical path and a detection optical path.

[0036] See in some examples Figure 2 and Figure 3 The illumination path may include: sleeve assembly 1, lens tube assembly 2, beam splitter 3, tube lens 4, and microscope objective lens 5.

[0037] In some examples, the microscope may include a first connecting device that enables the sleeve assembly 1 to move along the optical axis. The first connecting device can connect the sleeve assembly 1 to the microscope tube assembly 2, which together constitute a first module 100.

[0038] In some examples, the microscope may include a second connecting device that enables the tube mirror 4 to move along the optical axis. The second connecting device can connect the beam splitter 3 to the tube mirror 4, and the connection between the beam splitter 3 and the tube mirror 4 constitutes a second module 200.

[0039] In some examples, the first connecting device and the second connecting device may have a threaded structure, a slotted structure, or a guide rail structure.

[0040] See in some examples Figure 2 and Figure 3 The light can be emitted from the light source 11, enter the lens tube assembly 2 through the sleeve assembly 1, and then enter the spectrometer 3; the light passes through the spectrometer 3 and enters the tube lens 4, then enters the microscope objective lens 5, and finally the light shines on the surface of the sample being tested.

[0041] In some examples, the sleeve assembly 1 may include a light source 11, a collimating lens 12, and a field stop 13.

[0042] In some examples, the light source 11 may have a preset dark grid pattern, which can be used to determine whether the image of the light source 11 is clear, thus facilitating imaging observation in subsequent steps.

[0043] In some examples, the collimating lens 12 can effectively correct edge aberrations in the field of view and reduce spherical aberration, thereby improving light energy utilization.

[0044] In some examples, the field stop 13 may be positioned between the collimating lens 12 and the condenser lens 21 (described later).

[0045] In some examples, the sleeve assembly 1 may include an outer rotating cylinder 14, and the light source 11 may be located inside the outer rotating cylinder 14 of the sleeve assembly 1.

[0046] In some examples, the side wall of the outer rotating cylinder 14 of the sleeve assembly 1 may be provided with an adjustment device. In some examples, the adjustment device may be used to adjust the position of the light source 11. In some examples, the adjustment device may be used to adjust the position of the light source 11 in the vertical plane of the optical axis; in some examples, the adjustment device is a set screw.

[0047] In some examples, the sleeve assembly 1 may include an inner rotating cylinder 15. In some examples, the collimating lens 12 and the field stop 13 may be located inside the inner rotating cylinder 15 of the sleeve assembly 1.

[0048] In some examples, the light-gathering lens 21 may be placed in the first lens tube 24 of the lens tube assembly 2.

[0049] In some examples, the reflector 22 may be placed inside the second lens tube 25 of the lens tube assembly 2.

[0050] In some examples, the condenser lens 21 is also called a condenser or a condenser.

[0051] In some examples, the aperture stop 23 can be the through-hole portion of the second lens barrel 25.

[0052] In some examples, the light beam entering the lens tube assembly 2 can be focused by the light-collecting mirror 21 and reflected by the reflecting mirror 22 to the through-hole of the aperture stop 23.

[0053] In some examples, an observation window can be set at the aperture stop 23 to obtain the image of the light source (described later).

[0054] In some examples, the beam splitter 31 and the turntable 32 can be placed in the beam splitting chamber 3.

[0055] See in some examples Figure 1 The beam splitter 31 can be used to combine the measurement optical path and the illumination optical path.

[0056] In some examples, turntable 32 may be a turntable with a perforated surface. Specifically, turntable 32 may have perforations arranged in a certain pattern. In some examples, turntable 32 may be a Nipkow turntable.

[0057] In some examples, the portion with the small hole can also be referred to as the hole surface.

[0058] In some examples, in the illumination path, the beam entering the beam splitter 3 can reach the turntable 32 through the beam splitter 31, and then reach the tube mirror 4 through the small hole of the turntable 32.

[0059] In some examples, as described above, the image-side back focal plane and the parfocal plane of the microscope objective 5 can be a uniform fixed value.

[0060] In some examples, the spectrometer 3 may have a window or an openable side. In this case, the image on the turntable 32 can be observed visually or determined by instruments.

[0061] In some examples, after the light source 11 emits light that evenly illuminates the surface of the sample to be tested, the surface of the sample to be tested reflects the illumination light. The reflected light enters the microscope objective 5, then enters the tube mirror 4, the rotating disk 32, and the beam splitter 31. It undergoes a 90° reflection at the beam splitter 31 and then enters the detection optical path 6. Finally, the information of the sample to be tested is imaged on the imaging device of the detection optical path 6.

[0062] In some examples, the installation and adjustment method can install and adjust the lighting path in segments and zones. Figure 4A This is a flowchart illustrating a first embodiment of the assembly and adjustment method described in this example. Figure 4B This is a flowchart illustrating a second embodiment of the assembly and adjustment method described in this example.

[0063] See in some examples Figure 4A The setup and adjustment method may include: forming a first parallel beam (step S100); adjusting the image of the light source 11 on the aperture stop 23 (step S200); setting the turntable 32 and the beam splitter 31 in the beam splitter 3 (step S300); adjusting the tube lens 4 to focus the image (step S400); allowing the first parallel beam to enter the tube lens 4 (step S500); adjusting the image of the field stop 13 on the turntable 32 (step S600); determining the position of the microscope objective 5 and installing the microscope objective 5 (step S700). Based on the Köhler illumination requirements, the illumination condition of "pupil to window, window to pupil" is followed. The effect of this illumination condition is that the image of the light source 11 is formed on the image-side back focal plane of the microscope objective 5, and the field stop 13 and the object-side focal plane of the microscope objective 5 are conjugate. As a result, the surface of the illuminated object sample is smoothly and uniformly illuminated without shadows. By utilizing a reasonable illumination optical path structure and accurately adjusting the position of the observation window, the illumination optical path is finely calibrated step by step in sections, thereby improving the illumination accuracy of the light source 11 in the microscope imaging system and perfecting the imaging quality of the microscopic detection optical path.

[0064] See in some examples Figure 2 and Figure 8 The imaging process of light source 11 in the illumination optical path can be as follows: Figure 2 and Figure 8As shown by the solid line, the light source 11 forms its first image at the aperture stop 23. The second image is then placed on the image-side back focal plane of the microscope objective 5. In this case, the light source image, after passing through the microscope objective 5, is imaged at infinity on one side of the sample under test. Since the sample under test is at a finite distance, the surface illumination of the sample is not affected by the shadow of the light source filament. Therefore, the illumination light on the surface of the sample is uniform and there are no shadows. The imaging process of the field stop 13 in the illumination optical path can be found in [reference needed]. Figure 2 and Figure 8 As shown by the dashed line, the field stop 13 forms its image for the first time at the turntable 32. This image is the conjugate image of the field stop 13. At the same time, the field stop 13 and the object-side focal plane of the microscope objective 5 are conjugate. Thus, the field stop 13 determines the range of the illuminated object surface.

[0065] In some examples, the order of steps S100 and S200, and steps S300 and S400, can be changed. Specifically, steps S100 and S200 can precede steps S300 and S400; steps S100 and S200 can follow steps S300 and S400. See also [link to relevant documentation] for some examples. Figure 4B While executing steps S100 and S200 in sequence, steps S300 and S400 can be executed simultaneously.

[0066] Figure 5 This is a schematic diagram illustrating a scenario for acquiring a first parallel beam as described in this embodiment example.

[0067] In some examples, a first parallel beam can be formed in step S100. In this case, by collimating the beam of the light source 11, the edge rays can be focused and emitted in a parallel beam form, so that the light energy is uniformly distributed in the cross section perpendicular to the optical axis.

[0068] In some examples, the relative position between the light source 11 and the focal point of the collimating lens 12 can be changed by adjusting the position of the light source 11, so that the light source 11 is approximately located near the focal point of the collimating lens 12. In this case, the light source 11 forms a first parallel beam through the collimating lens 12, and the first parallel beam passes through the field stop 13 and enters the lens barrel assembly 2. In this case, the collimating lens 12 can effectively correct the aberrations at the edge of the field of view, reduce spherical aberration, and improve the light energy utilization rate. At the same time, the light is emitted in the form of a parallel beam, so that the light energy is uniformly distributed in the cross section perpendicular to the optical axis, and the shape of the light source 11 is imaged at infinity.

[0069] In some examples, the initial position of light source 11 is near the theoretical focal length of collimating lens 12. (Reference) Figure 5An observation window is set on the outer side of the inner rotating cylinder 15, away from the light source 11. By moving the observation window along the optical axis, the shape and size of the light spot on the observation window are observed. If the outer diameter of the light spot changes with the movement of the observation window, the outer rotating cylinder 14 needs to be further adjusted, thereby adjusting the focal position of the light source 11 on the collimating lens 12, until the shape and size of the light spot remain consistent at different selected positions, and the outer diameter of the light spot does not change with the movement of the observation window, maintaining a consistent state. In some examples, 3-5 positions can be selected to place the observation window to observe the shape and size of the light spot. Then, the outer rotating cylinder 14 is fixed, the light source collimation is completed, and step S100 is finished. It should be noted that "the shape and size of the light spot remain consistent" can be understood in the following ways: the shape and size of the light spot are determined by visual inspection, and the consistency of the light spot shape and size is determined based on the visual inspection; or the shape and size of the light spot are obtained by instruments and equipment, and the consistency of the light spot shape and size is determined based on preset rules.

[0070] In some examples, after adjusting the relative positions of the light source 11 and the collimating lens 12, the positional relationship between the light source 11 and the collimating lens 12 is fixed. In this case, partitioned assembly and adjustment can be achieved, ensuring that other assembly and adjustment steps do not affect the relative positional relationship between the assembled modules.

[0071] In some examples, adjustments can be made manually. In others, as the observation window moves, changes in the outer diameter of the light spot can be judged by visual observation of the window. In these cases, adjustments can be made conveniently.

[0072] In some examples, digital imaging observation technology can be used to achieve automatic software calibration. The digital imaging observation system includes a scanning / control unit, an image acquisition / processing unit, and a computer-aided measurement / control unit.

[0073] In some examples, multiple observation windows can be set simultaneously, and it can be determined whether the outer diameter shape and size of the light spot in the multiple observation windows are consistent, thereby determining whether the light beam passing through the collimating lens 12 is collimated.

[0074] In some examples, the relative position of the light source 11 and the collimating lens 12 can be adjusted to form a first parallel beam of light incident on the lens barrel assembly 2 via the collimating lens 12. In some examples, the collimating lens 12 uses a group of spherical mirrors to collimate the light from the light source.

[0075] In some examples, the light beam from the light source 11 in the illumination path is collimated by the collimating lens 12 and emitted as parallel light. Near the collimating lens 12, the light rays in the cross section of the optical axis are uniformly distributed. It is suitable to set the field stop 13 here. Setting the field stop 13 here can effectively limit the light beam, thereby effectively limiting the imaging range of the light source 11.

[0076] In some examples, the field stop 13 limits the field of view of the optical path system, determining the size of the illuminated object surface. The aperture value of the field stop 13 is a preset value in the optical design of the microscope illumination path.

[0077] In some examples, as described above, the field stop 13 can be positioned between the collimating lens 12 and the condenser lens 21.

[0078] In some examples, the field stop 13 can be set outside one focal length and inside two focal lengths on the light source side of the condenser lens 21. In this case, the conjugate image of the field stop 13 through the condenser lens 21 is presented on the turntable 32. Its conjugate image is a magnified real image with uniform light distribution in the image plane, which meets the illumination requirements of the turntable 32. At the same time, the field stop 13 and the object-side focal plane of the microscope objective 5 are a pair of conjugate planes, thereby ensuring that the illumination light on the surface of the illuminated object sample is smooth and uniform without shadows.

[0079] In some examples, the field stop 13 has a non-circular through-hole. In some examples, the field stop 13 is square in shape. In this case, it is easy to observe the shape of the field stop 13 on the small hole surface of the turntable 32 during the assembly process. However, the through-hole of the field stop 13 is not limited to square. For example, the through-hole of the field stop 13 can be star-shaped or irregular in shape. In some cases, since the outer shell of the lens assembly 2 or the frame of other components (such as the outer frame of the lens) may also serve as the actual effective field stop, and in the optical path system, the shape of the outer shell of the first lens 24 or the frame of other components is generally circular, rectangular, polygonal or elliptical, the field stop 13 with a non-circular through-hole can be used to determine that the field stop 13 provided on the sleeve assembly 1 is the one that limits the beam. In other words, compared to using the outer frame of a lens as a field stop or omitting the field stop, this disclosure makes it easier for the adjuster to distinguish the image of the field stop 13 during optical path adjustment by using a field stop 13 with a non-circular through-hole, thus achieving accurate adjustment.

[0080] In some examples, the field stop 13 is a variable aperture stop, which can be adapted to different optical design requirements. The microscope can be replaced with a variable aperture stop of different diameters according to the optical specifications.

[0081] Figure 6 This is a schematic diagram illustrating a scene involving adjusting the posture of a light source as described in this embodiment example.

[0082] In some examples, step S200 may be performed after step S100.

[0083] In some examples, in step S200, the imaging of the light source 11 onto the aperture stop 23 can be adjusted. In this case, adjusting the orientation of the light source 11 so that it forms a centrally symmetrical image in the aperture stop 23 enables the optical axis of the parallel beam to pass through the center of the aperture stop 23, and allows the orientation of the light source 11 to be adjusted so that there is no loss of illumination energy, thereby reducing aberrations caused by the optical axis not passing through the center of the aperture stop 23.

[0084] In some examples, reference Figure 3 and Figure 6 The lens barrel assembly 2 may include a first lens barrel 24 and a second lens barrel 25.

[0085] In some examples, the condenser lens 21 can be placed inside the first lens barrel 24, and the reflector 22 can be placed inside the second lens barrel 25; the aperture stop 23 is a through-hole portion of the second lens barrel 25. A first connecting device can connect the sleeve assembly 1 to the lens barrel assembly 2, and the first lens barrel 24 can be moved along the optical axis via the first connecting device. Light from the sleeve assembly 1 passes through the condenser lens 21, illuminates the reflector 22, is reflected by the reflector 22 to the aperture stop 23, and then passes through the aperture stop 23.

[0086] In some examples, in a microscopic illumination system, a condenser lens 21 can be placed after the collimating optical path of the light source (i.e., the optical path that forms the first parallel beam in step S100). In this case, the condenser lens 21 increases the energy density of the illumination beam, giving the microscope high resolution and contrast characteristics, and improving the illumination light energy.

[0087] In some examples, the sleeve assembly 1 and the first lens barrel 24 of the lens barrel assembly 2 can be connected. In some examples, the sleeve assembly 1 and the lens barrel assembly 2 are connected by threads. In this case, the sleeve assembly 1 can move along the optical axis, thereby adjusting the relative position of the sleeve assembly 1 and the lens barrel assembly 2.

[0088] In some examples, an observation window can be provided at the through-hole of the aperture stop 23. In this case, the image of the light source 11 at the aperture stop 23 can be observed through the observation window. In other words, the observation window can be provided at the aperture stop 23 to receive the image of the light source 11 formed by the aperture stop 23. Since it is difficult to obtain the image of the light source when the light beam passes through the through-hole of the aperture stop 23, by using the observation window provided at the aperture stop 23, the light beam passing through the through-hole of the aperture stop 23 can be obtained to form the image of the light source, and thus the relative position of the aperture stop 23 and the condenser lens 21 can be determined.

[0089] In some examples, the aperture stop 23 can be initially positioned near the focal plane of the condenser lens 21 to control the aperture angle of the illumination system. The light beam from the light source 11 passes through the collimating lens 12 to form a first parallel beam, which then passes through the condenser lens 21 and is imaged at the aperture stop 23.

[0090] In some examples, the image of the light source 11 on the observation window, which is located at the through-hole of the aperture stop 23, can be observed by moving the first lens tube 24 along the optical axis. In this embodiment, the light source 11 may have a preset dark grid pattern. When a preset clear image of the light source wick dark pattern appears in the observation window, the first lens tube 24 can be fixed. This ensures that the aperture stop 23 is located at the focal plane of the condenser lens 21, thereby ensuring that the aperture stop 23, the condenser lens 21, and the collimating lens 12 are on the same optical axis and placed perpendicular to the optical axis without tilt, thus preventing aberrations in the image of the light source.

[0091] In some examples, after adjusting the relative positions of the field stop 23 and the condenser lens 21, their positional relationship is fixed. In this case, partitioned assembly and adjustment can be achieved, ensuring that other assembly and adjustment steps do not affect the relative positional relationship between the assembled modules.

[0092] In some examples, after determining that the aperture stop 23 is located at the focal plane of the condenser lens 21, the orientation of the light source 11 can be adjusted so that the light source 11 forms a centrally symmetrical image in the aperture stop 23. In some examples, the side wall of the outer rotating cylinder 14 of the sleeve assembly 1 is provided with an adjustment device, which can be used to adjust the position of the light source 11 in the vertical plane of the optical axis. In some examples, the adjustment device is a set screw. Thus, by using this adjustment device, the orientation of the light source 11 can be adjusted so that the light source 11 forms a centrally symmetrical image in the aperture stop. Furthermore, by adjusting the orientation of the light source 11, the position of the light source 11 can be adjusted up, down, left, and right in the plane perpendicular to the optical axis, while observing the observation window, so that the image of the light source 11 is completely symmetrical within the aperture stop 23, and the edge of the light source image is completely within the aperture range of the aperture stop 23, thereby ensuring no loss of illumination energy.

[0093] In some examples, the observation window in step S200 can be a semi-transparent object; in others, it can be a semi-transparent frosted glass, which facilitates visual observation. Simultaneously, the image formed by the light source 11 at the aperture stop 23 is projected onto the semi-transparent object, thus facilitating visual observation of the imaging.

[0094] In some examples, step S300 may be performed after step S200.

[0095] In some examples, in step S300, the turntable 32 and the beam splitter 31 can be placed in the beam splitting chamber 3, and their positional distribution can be preset values ​​for the optical design.

[0096] In some examples, as described above, the turntable 32 may have many small holes distributed on it, and the light beam of the light source 11 covers the range of all the small holes (i.e., the scanning area). When the turntable 32 rotates at high speed, one small hole scans a corresponding area on the sample to be tested, thereby achieving a complete scan of the sample to be tested.

[0097] In some examples, step S400 may be performed after step S300.

[0098] In some examples, in step S400, the scope 4 can be adjusted to focus the image. Figure 7 This diagram illustrates the relative positions of the adjusting lens 4 and the beam splitter 3 as described in this embodiment example.

[0099] See in some examples Figure 7 The tube mirror 4 can be connected to the spectrometer 3.

[0100] In some examples, after the lens 4 is connected to the beam splitter 3, a second parallel beam can be formed and directed from the lens 4 into the beam splitter 3.

[0101] In some examples, the second parallel beam can be generated by the collimator 7, which has a preset image, and the second parallel beam is directed from the lens 4 into the beam splitter 3. The relative positions of the lens 4 and the beam splitter 3 are adjusted so that the parallel beam forms a preset image on the turntable 32.

[0102] In some examples, the preset image can be a collimator reticle image that matches the collimator 7. Since the reticle image is imaged at infinity through the second parallel beam, the collimator 7 can simulate the reflected beam of the sample being tested passing through the microscope objective 5 and entering the turntable 32, thereby enabling more accurate adjustment of the relative position of the collimator 4 and the spectrometer 3.

[0103] In some examples, the lens 4 and the beam splitter 3 are connected by threads. In this case, the lens 4 and the beam splitter 3 can be rotated along the threads, causing the lens 4 to move along the optical axis. This allows adjustment of the relative position between the lens 4 and the beam splitter 3, thereby adjusting the relative position between the lens 4 and the turntable 32.

[0104] In some examples, the entire assembly of the microscope tube 4 and the beam splitter 3 is positioned under the collimator 7. The collimator 7 forms a second parallel beam that enters the beam splitter 3 from the microscope tube 4. The relative positions of the microscope tube 4 and the beam splitter 3 are adjusted so that the second parallel beam produces a clear image of the reticle of the collimator 7 on the rotating disk 32. In this case, by simulating the reflection of illumination light from the surface of the sample under test, the reflected light enters the microscope objective 5, enters the microscope tube 4 as a parallel beam, and forms an image on the rotating disk 32. This allows the relative positions of the microscope tube 4 and the rotating disk 32 to be determined, ensuring the integrity of the optical path.

[0105] In some examples, after adjusting the relative positions of the lens 4 and the beam splitter 3, the positional relationship between the lens 4 and the beam splitter 3 is fixed. In this case, partitioned assembly and adjustment can be achieved, ensuring that other assembly and adjustment steps do not affect the relative positional relationship between the assembled modules.

[0106] In some examples, step S500 may be performed after step S400.

[0107] In some examples, in step S500, the first parallel beam can be directed into the tube lens 4. Specifically, the first module 100 can be placed on the second module 200, and the second module 200 is placed on the side of the condenser lens 21 away from the light source 11, at a position greater than twice the focal length of the condenser lens 21, so that the first parallel beam is directed into the beam splitter 3 through the sleeve assembly 1 and the lens barrel assembly 2, and then enters the tube lens 4 through the beam splitter 3.

[0108] In some examples, step S600 may be performed after step S500.

[0109] In some examples, in step S600, the imaging of the field stop 13 on the turntable 32 can be adjusted. In this case, by adjusting the relative positions of the field stop 13, the turntable 32, and the microscope objective 4 in segments and sections, and making the field stop 13 form an image of the field stop 13 on the aperture surface of the turntable 32, and the image of the field stop 13 covers the aperture surface of the turntable, it is possible to achieve conjugate between the field stop 13 and the object-side focal plane of the microscope objective 5.

[0110] In some examples, an infinite conjugate image microscope objective 5 and a tube lens 4 can be used together to achieve microscopic imaging. In a rotary parallel scanning confocal microscope, the aperture on the rotary disk 32 is shared by the illumination and detection imaging paths. The image plane, i.e., the conjugate image plane of the object plane, lies on the aperture surface of the rotary disk 32. Therefore, the image of the field stop 13 needs to be presented on the aperture surface of the rotary disk 32, and the light spot of the field stop 13 image plane completely covers the aperture surface within the imaging range of the rotary disk 32. It should be noted that the imaging range can be understood as follows: during the operation of the parallel scanning confocal microscope, the rotary disk 32 rotates along its central axis, and a portion of the rotary disk 32 lies within both the illumination and measurement paths. The area on the rotary disk 32 located within the illumination and measurement paths can be called the imaging range.

[0111] In some examples, the position of the sleeve assembly 1 can be adjusted by moving it along the optical axis. In this case, the light source 11, collimating lens group 12, and field stop 13 are moved as a single module relative to the lens barrel assembly 2 along the optical axis, thereby adjusting the relative position of the field stop 13 and the condenser lens 21 so that a clear image of the field stop 13 is formed on the turntable 32, and the light spot of the image plane of the field stop 13 completely covers the pinhole surface within the imaging range of the turntable 32. See [reference needed]. Figure 2 and Figure 8 As shown, the dashed line in the figure represents the imaging process of the field stop 13. In some examples, as described above, the beam splitter 3 may have an openable side, in which case the image spot of the field stop 13 on the turntable 32 can be observed.

[0112] In some examples, the actual field stop 13 image plane spot size is the illumination path design value. The illumination path design value can be calculated using the Gaussian formula.

[0113] In some examples, step S700 can be performed after step S600. In some examples, in step S700, the position of the microscope objective 5 can be determined and the microscope objective 5 can be installed. In this case, by placing an observation window on the side of the tube mirror 4 away from the beam splitter 3, the observation window is observed until it receives an image of the light source 11 that meets the preset requirements. At this time, the position of the observation window is the image-side back focal plane of the microscope objective 5. Through the design parameters of the microscope objective 5, the numerical relationship between the image-side back focal plane and the parfocal plane of the microscope objective 5 can be determined, and thus the installation position of the microscope objective 5 can be determined, thereby enabling the light source to be imaged on the image-side back focal plane of the microscope objective 5.

[0114] Figure 8 This is a schematic diagram illustrating a scenario involving adjusting the position of a microscope objective as described in this embodiment example.

[0115] In some examples, before assembling the microscope objective 5 in the overall optical path, it is necessary to first locate a clear image of the light source formed by the beam of the light source 11 after passing through the sleeve assembly 1, the lens tube assembly 2, the beam splitter 3, and the tube lens 4, and mark the position of this clear image. The image-side back focal plane of the microscope objective 5 is then set at this position. In this case, the image of the light source is imaged at infinity on one side of the sample being tested after passing through the microscope objective 5. Since the sample being tested is at a finite distance, the illumination of the sample surface is not affected by the shadow of the light source filament. Therefore, the illumination light on the sample surface is uniform and without shadows. In other words, see... Figure 2 and Figure 8 As shown by the solid line, the light source 11 forms a first real image at the aperture stop 23 through the collimating lens 12 and the condenser lens 21. The first image then forms a second real image between the tube lens 4 and the microscope objective 5 after passing through the tube lens 4. Thus, the microscope objective 5 can be assembled based on the positions of the first and second real images.

[0116] In some examples, reference Figure 8 An observation window can be placed on the side of the tube lens 4 away from the beam splitter 3, and the observation window can be moved along the optical axis to receive the image of the light source 11 formed by the beam emitted from the tube lens 4 at different positions until a clear image of the light source 11 that meets the preset requirements appears on the observation window, and the position of the observation window at this time is marked.

[0117] In some examples, the image of the light source 11 formed by the beam emitted from the tube lens 4 can be received through the observation window. The observation window is moved along the optical axis until an image of the light source 11 that meets the preset requirements appears on the observation window. This allows the position of the clear image of the light source 11 to be determined, and consequently, the mounting position of the image-side back focal plane of the microscope objective 5 can be determined. Thus, the mounting position of the microscope objective 5 can be determined based on the numerical relationship between the image-side back focal plane and the parfocal plane.

[0118] In some examples, the preset requirement may be that the image of light source 11 has a certain sharpness and shape. In some examples, the preset requirement may be that the operator can clearly distinguish the shape of the image of light source 11. In some examples, whether the image of light source 11 meets the preset requirement may be determined by image processing methods; for example, the preset requirement may be that the resolution (or gradient) of the image of light source 11 meets the requirement.

[0119] In some examples, the image-side back focal plane of the microscope objective 5 can be set near or at this location. This location is where the image-side back focal plane of the microscope objective 5 is placed.

[0120] In some examples, as described above, the image-side back focal plane of the microscope objective 5 and the parfocal plane of the microscope objective 5 are a unified fixed value. In this case, the mounting position of the microscope objective 5 can be determined based on the unified fixed value of the image-side back focal plane of the microscope objective 5 and the parfocal plane of the microscope objective 5.

[0121] In some examples, as described above, the image-side back focal plane and parfocal plane of the microscope objective 5 can be a uniform fixed value. This allows for the determination of the mounting position of the microscope objective 5 and its installation.

[0122] In some examples, the observation window in step S700 above is a semi-transparent object; in other examples, the semi-transparent object is semi-transparent frosted glass. In this case, it is convenient for visual observation. At the same time, the image of the light source 11 formed by the light beam emitted from the tube lens 4 is imaged on the semi-transparent object, thereby facilitating visual observation of the corresponding imaging situation.

[0123] Various embodiments of the invention have been described above in detail. Although these descriptions directly depict the above embodiments, it should be understood that modifications and / or variations to the specific embodiments shown and described herein will occur to those skilled in the art. Any such modifications or variations falling within the scope of this specification are also intended to be included herein. Unless specifically indicated, the inventors intend that the words and phrases in the specification and claims be given the common and customary meaning to those skilled in the art.

[0124] The above description of various embodiments of the invention known to the applicant at the time of filing this application is intended for illustrative and descriptive purposes. This description is not intended to be exhaustive, nor does it limit the invention to the exact forms disclosed, and many modifications and variations can be made based on the foregoing teachings. The described embodiments are intended to explain the principles of the invention and its practical application, and to enable others skilled in the art to utilize the invention in various embodiments and with various modifications suitable for the intended particular use. Therefore, the invention is not intended to be limited to the specific embodiments disclosed for implementing the invention.

[0125] While specific embodiments of the invention have been shown and described, it will be apparent to those skilled in the art that variations and modifications can be made based on the teachings of the invention without departing from the invention and its broader aspects, and therefore the appended claims are intended to cover all such changes and modifications within the true spirit and scope of the invention. Those skilled in the art will understand that, in general, the terminology used in this invention is intended to be “open” terminology (e.g., the term “comprising” should be interpreted as “including but not limited to”, the term “having” should be interpreted as “at least having”, the term “comprising” should be interpreted as “including but not limited to”, etc.).

Claims

1. An illumination optical path for a rotary parallel scanning confocal microscope, characterized in that, The illumination optical path includes a sleeve assembly with a light source and a field stop, a microscope tube assembly with a condenser lens, a beam splitter with the turntable, a tube mirror, and a microscope objective. The light source is configured to form a first parallel beam entering the microscope tube assembly via the field stop; the light source is imaged at the focal plane of the condenser lens; the turntable is configured to receive a second parallel beam incident from the tube mirror, the second parallel beam being configured to form a preset image on the turntable; the turntable is also configured to receive the first parallel beam incident from the microscope tube assembly into the beam splitter, the first parallel beam being configured to form a spot covering a small aperture surface of the turntable; the microscope objective is configured to be mounted at the position of the image of the light source formed by the beam emitted from the tube mirror, conforming to preset requirements; the microscope tube assembly also includes an aperture stop, in which the aperture stop is positioned... An observation window is provided at the aperture stop. The lens assembly includes a first lens tube and a second lens tube. The condenser lens is disposed inside the first lens tube. The aperture stop is the through-hole portion of the second lens tube. The illumination optical path also includes a first connecting device that connects the sleeve assembly to the lens tube assembly. The first connecting device is configured to move the first lens tube along the optical axis. The observation window is configured to observe the image of the light source at the aperture stop so that the aperture stop is positioned on the focal plane of the condenser lens via the first connecting device. The side wall of the outer rotating cylinder of the sleeve assembly is provided with an adjustment device for adjusting the position of the light source in the vertical plane of the optical axis. The observation window is also configured to observe the image of the light source at the aperture stop so that the attitude of the light source is adjusted via the adjustment device so that the light source forms a centrally symmetrical image in the aperture stop.

2. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 1, characterized in that, Adjust the relative position of the tube mirror and the beam splitter so that the second parallel beam forms a preset image on the turntable.

3. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 2, characterized in that, The first parallel beam is directed into the beam splitter through the lens assembly and forms a light spot on the turntable. The relative positions of the field stop and the condenser lens are adjusted to form a light spot covering the small aperture surface of the turntable.

4. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 3, characterized in that, The sleeve assembly includes a collimating lens, and the relative position of the light source and the collimating lens is adjusted to form the first parallel beam in the collimating lens.

5. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 4, characterized in that, After adjusting the relative positions of the light source and the collimating lens, fix the positional relationship between the light source and the collimating lens; after adjusting the relative positions of the field stop and the condenser lens, fix the positional relationship between the field stop and the condenser lens; after adjusting the relative positions of the tube mirror and the beam splitter, fix the positional relationship between the tube mirror and the beam splitter.

6. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 1, characterized in that, The image of the light source formed by the light beam emitted from the tube lens is received at different positions to obtain the position of the image of the light source that meets the preset requirements.

7. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 1 or 6, characterized in that, The image of the light source formed by the light beam emitted from the tube lens is received through the observation window. The observation window is moved along the optical axis until the image of the light source that meets the preset requirements appears on the observation window.

8. The illumination optical path of the rotary parallel scanning confocal microscope according to claim 1, characterized in that, The field stop has a non-circular through-hole.

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