CN220572188U - Multiphoton blood flow microscopic imaging system - Google Patents
Multiphoton blood flow microscopic imaging system Download PDFInfo
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- CN220572188U CN220572188U CN202321685828.6U CN202321685828U CN220572188U CN 220572188 U CN220572188 U CN 220572188U CN 202321685828 U CN202321685828 U CN 202321685828U CN 220572188 U CN220572188 U CN 220572188U
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- 230000017531 blood circulation Effects 0.000 title claims abstract description 39
- 238000003384 imaging method Methods 0.000 title claims abstract description 33
- 238000001514 detection method Methods 0.000 claims abstract description 33
- 230000005284 excitation Effects 0.000 claims abstract description 14
- 230000010287 polarization Effects 0.000 claims abstract description 10
- 230000003287 optical effect Effects 0.000 claims description 34
- 238000012545 processing Methods 0.000 claims description 5
- 238000000386 microscopy Methods 0.000 claims description 2
- 210000004204 blood vessel Anatomy 0.000 abstract description 15
- 230000000007 visual effect Effects 0.000 abstract description 5
- 238000011160 research Methods 0.000 abstract description 4
- 230000008859 change Effects 0.000 description 4
- 238000010586 diagram Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 230000006641 stabilisation Effects 0.000 description 2
- 238000011105 stabilization Methods 0.000 description 2
- 230000003321 amplification Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 238000000034 method Methods 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 239000013307 optical fiber Substances 0.000 description 1
- 239000005304 optical glass Substances 0.000 description 1
- 238000012634 optical imaging Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
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Abstract
The multi-photon blood flow microscopic imaging system provided by the embodiment of the utility model comprises a first laser, a two-dimensional scanning unit, a first dichroic mirror, a zoom objective lens, a first signal detection unit, a second laser, a second dichroic mirror, a polarization spectroscope, a second signal detection unit and a control unit; the two-dimensional scanning unit is arranged at one side of the first dichroic mirror, which is away from the zoom objective lens; the excitation light emitted by the first laser irradiates the sample through the first dichroic mirror and the zoom objective lens, and fluorescence emitted after the sample is excited passes through the zoom objective lens and is reflected at the first dichroic mirror to enter the first signal detection unit; after being reflected by the second dichroic mirror, the laser emitted by the second laser irradiates the sample through the zoom objective to scatter, scattered light passes through the zoom objective and is reflected into the second signal detection unit at the polarization spectroscope, and the speckle image of blood flow and the three-dimensional image of blood vessel distribution are synthesized, so that the observation and research on tiny blood vessels are more accurate and visual.
Description
Technical Field
The utility model relates to the technical field of optical imaging, in particular to a multiphoton blood flow microscopic imaging system.
Background
Laser speckle blood flow imaging is a regional flow velocity monitoring technology, and living organisms are subjected to living body, dynamic and non-contact monitoring on blood flow velocity, vessel diameter, blood flow change and the like under the condition of no need of scanning, so that a plurality of indexes of blood flow dynamics are obtained.
However, the speckle image is a two-dimensional image from which depth information of blood vessels cannot be acquired. If the tiny blood vessels are staggered up and down, only the blood flow diagrams which are intersected with each other can be seen on the speckle image, and the three-dimensional space distribution of the blood vessels cannot be seen, so that the observation is not accurate and visual enough.
Disclosure of Invention
The embodiment of the utility model provides a multiphoton blood flow microscopic imaging system, which aims to solve the problem that blood flow can be observed only through a two-dimensional image in the prior art.
In a first aspect, there is provided a multiphoton blood flow microscopy imaging system comprising:
the device comprises a first laser (1), a two-dimensional scanning unit (2), a first dichroic mirror (3), a zoom objective lens (4), a first signal detection unit (5), a second laser (11), a second dichroic mirror (16), a polarization spectroscope (13), a second signal detection unit (15) and a control unit (6);
the first laser (1), the two-dimensional scanning unit (2), the zoom objective lens (4), the first signal detection unit (5), the second laser (11) and the second signal detection unit (15) are respectively and electrically connected with the control unit (6);
the two-dimensional scanning unit (2) is arranged at one side of the first dichroic mirror (3) away from the zoom objective lens (4);
the optical axis of the first signal detection unit (5) is perpendicular to the optical axis of the zoom objective lens (4) and intersects the first dichroic mirror (3);
the optical axis of the second laser (11) is perpendicular to the optical axis of the zoom objective (4) and intersects the second dichroic mirror (16);
the optical axis of the second signal detection unit (15) is perpendicular to the optical axis of the zoom objective lens (4) and is intersected with the polarization beam splitter (13);
after passing through the two-dimensional scanning unit (2), the excitation light emitted by the first laser (1) irradiates a sample through the first dichroic mirror (3) and the zoom objective (4), and fluorescence emitted by the excited sample transmits through the zoom objective (4) and is reflected at the first dichroic mirror (3) to enter the first signal detection unit (5);
after being reflected by a second dichroic mirror (16), the laser emitted by the second laser (11) irradiates the sample through the zoom objective (4) to be scattered, and the scattered light passes through the zoom objective (4) and is reflected at a polarization spectroscope (13) to enter a second signal detection unit (15).
According to the embodiment of the utility model, the speckle image of blood flow is obtained through laser speckle blood flow imaging, the three-dimensional image of blood vessel distribution is obtained through multiphoton microscopic imaging, and the speckle image of blood flow and the three-dimensional image of blood vessel distribution are synthesized, so that the observation and research on tiny blood vessels are more accurate and visual.
Drawings
The foregoing and/or additional aspects and advantages of the utility model will become apparent and readily appreciated from the following description of the embodiments, taken in conjunction with the accompanying drawings, in which:
fig. 1 is a schematic diagram of a multiphoton blood flow microscopic imaging system according to an embodiment of the present utility model.
Description of the embodiments
Embodiments of the present utility model are described in detail below, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to like or similar modules or modules having like or similar functions throughout. The embodiments described below by referring to the drawings are illustrative only and are not to be construed as limiting the utility model. On the contrary, the embodiments of the utility model include all alternatives, modifications and equivalents as may be included within the spirit and scope of the appended claims.
It should be noted that all directional indicators (such as up, down, left, right, front, and rear) in the embodiments of the present utility model are merely used to explain the relative positional relationship, movement, etc. between the components in a specific posture. If the particular gesture changes, the directional indication changes accordingly.
It will be further understood that when an element is referred to as being "mounted" or "disposed" on another element, it can be directly on the other element or intervening elements may also be present. When an element is referred to as being "connected" to another element, it can be directly connected to the other element or be indirectly connected to the other element through intervening elements.
According to the embodiment of the utility model, the speckle image of blood flow is obtained through laser speckle blood flow imaging, the three-dimensional image of blood vessel distribution is obtained through multiphoton microscopic imaging, and the speckle image of blood flow and the three-dimensional image of blood vessel distribution are synthesized, so that the observation and research on tiny blood vessels are more accurate and visual.
Fig. 1 is a schematic diagram of a multiphoton blood flow microscopic imaging system according to an embodiment of the present utility model. As shown in fig. 1, the multiphoton blood flow microscopic imaging system includes a first laser 1, a two-dimensional scanning unit 2, a first dichroic mirror 3, a zoom objective lens 4, a first signal detection unit 5, a second laser 11, a second dichroic mirror 16, a polarizing beam splitter 13, a second signal detection unit 15, and a control unit 6. The first laser 1, the two-dimensional scanning unit 2, the zoom objective lens 4, the first signal detection unit 5, the second laser 11 and the second signal detection unit 15 are respectively electrically connected with the control unit 6, and controlled by the control unit 6. The control unit 6 may be a personal computer (Personal Computer, PC), a microcomputer, or the like.
In the embodiment of the present utility model, the first laser 1 is an optical fiber femtosecond laser, and is an excitation light source for multiphoton microscopic imaging, and the center wavelength of the emitted excitation light includes, but is not limited to 808 nm, 970 nm, 1310 nm, and the like.
The two-dimensional scanning unit 2 is arranged on the side of the first dichroic mirror 3 facing away from the zoom objective 4, and comprises a first galvanometer 21 and a second galvanometer 22, which are electrically connected to the control unit 6, respectively, the first galvanometer 21 and the second galvanometer 22 being arranged in an orthogonal manner. The first galvanometer mirror 21 and the second galvanometer mirror 22 may be galvanometer mirrors or resonant mirrors. The specific program of the control unit 6 regularly changes the reflection angle of the first galvanometer 21 to scan in the X-axis direction and changes the reflection angle of the second galvanometer 22 to scan in the Y-axis direction, thereby realizing two-dimensional scanning of the sample.
The first dichroic mirror 3 is disposed at an angle of 45 degrees with respect to the optical axis of the zoom objective lens 4, and is capable of transmitting excitation light emitted from the first laser 1 and reflecting fluorescence, and is used for splitting the excitation light and the fluorescence.
The zoom objective 4 includes a liquid lens 41 and other optical glass lenses electrically connected to the control unit 6. The control unit 6 changes the focal length of the liquid lens 41 by changing the electrical parameter. The focal length of the zoom objective lens 4 changes, and the focal point of the zoom objective lens in the Z-axis direction also changes, so that the scanning in the Z-axis direction is realized. The Z-axis scanning is combined with the X-axis scanning and the Y-axis scanning, so that three-dimensional imaging of the sample is realized.
The optical axis of the first signal detection unit 5 is perpendicular to the optical axis of the zoom objective lens 4, intersects the first dichroic mirror 3, and includes a converging lens 51 and a photomultiplier 52 coaxially disposed. The condensing lens 51 collects as much fluorescence as possible to be condensed on the photosensitive surface of the photomultiplier tube 52. After the photomultiplier tube 52 acquires weak fluorescence, it converts the weak fluorescence into an electrical signal and transmits the electrical signal to the control unit 6. Preferably, the first signal detection unit 5 further includes a first filter 53 coaxially disposed with the converging lens 51. The first optical filter 53 is disposed on a side of the converging lens 51 away from the photomultiplier 52, and prevents the excitation light from entering the photomultiplier 52, thereby improving the signal-to-noise ratio.
In the embodiment of the present utility model, the second laser 11 is a frequency stabilized laser, and outputs stable single-mode laser, and its center wavelength is 785 nm, for example, a near infrared laser source. The second laser 11 comprises a frequency stabilized laser diode, a constant power driver and a constant temperature controller. The constant power driver enables the frequency stabilization laser diode to output stable laser power, and the constant temperature controller enables the light emitting chip of the frequency stabilization laser diode to work at a constant temperature, so that the second laser 11 is ensured to have stable optical properties. The second laser 11 is a light source for laser speckle blood imaging, the optical axis of which is perpendicular to the optical axis of the zoom objective 4 and intersects the second dichroic mirror 16. Preferably, a laser collimating mirror 12 is provided between the second laser 11 and the second dichroic mirror 16. The laser collimator lens 12 is composed of a plurality of lenses or a single spherical mirror for collimating the laser light emitted from the second laser 11.
The first dichroic mirror 3, the second dichroic mirror 16 and the polarizing beam splitter 13 are arranged along the optical axis of the zoom objective 4, the second dichroic mirror 16 being arranged on the side of the polarizing beam splitter 13 facing away from the first dichroic mirror 3. The second dichroic mirror 16 and the polarizing beam splitter 13 are arranged on the side of the first dichroic mirror 3 facing away from the zoom objective 4. As a result, the first dichroic mirror 3 can transmit the laser light emitted from the second laser 11 in addition to the excitation light and the reflected fluorescence emitted from the first laser 1, and can transmit the scattered light in the reverse direction. The second dichroic mirror 16 is disposed at an angle of 45 degrees with respect to the optical axis of the zoom objective lens 4, and is capable of reflecting the laser light emitted from the second laser 11 and transmitting the excitation light emitted from the first laser 1. The polarizing beam splitter 13 is disposed at an angle of 45 degrees with respect to the optical axis of the zoom objective lens 4, and is capable of transmitting the excitation light emitted from the first laser 1, the laser light emitted from the second laser 2, and the reflected and scattered light.
The optical axis of the second signal detection unit 15 is perpendicular to the optical axis of the zoom objective 4, intersecting the polarizing beam splitter 13. The second signal detection unit 15 may be a photo array sensor. After the scattered light is acquired by the photo array sensor, it is converted into an electrical signal and transmitted to the control unit 6. Preferably, a second optical filter 14 is disposed between the second signal detecting unit 15 and the polarizing beam splitter 13 to prevent light other than scattered light from entering the photoelectric array sensor, thereby improving the signal-to-noise ratio.
The sample is placed on the focal plane of one side of the zoom objective lens 4 far away from the first dichroic mirror 3, after the excitation light emitted by the first laser 1 is scanned in the X-axis direction and the Y-axis direction by the two-dimensional scanning unit 2, the sample is irradiated through the first dichroic mirror 3 and the zoom objective lens 4, and the photons (fluorescence) emitted after the sample is excited and a small amount of the excitation light scattered by the sample reversely penetrate through the zoom objective lens 4, and are reflected at the first dichroic mirror 3 to enter the first signal detection unit 5. The excitation light is intercepted by the first filter 53 from reaching the photomultiplier tube 52, the fluorescence passes through the first filter 53 and the condensing lens 51 to reach the photomultiplier tube 52, is converted into an electric signal, and is transmitted to the PC, and then the three-dimensional image of the sample is reconstructed by the PC.
After being reflected by the second dichroic mirror 16, the laser light emitted by the second laser 11 irradiates the sample through the polarizing beam splitter 13, the first dichroic mirror 3 and the zoom objective lens 4, and the laser light penetrates into the sample to a certain depth, and is absorbed, reflected, transmitted, scattered, and the like therein. In this process, the polarization characteristics of the light change, and the polarization state of the scattered light is different from that of the incident light. The scattered light is back transmitted through the zoom objective 4 and the first dichroic mirror 3, reflected at the polarizing beam splitter 13 into the second signal detection unit 15. Light other than the scattered light is intercepted by the second filter 14, the scattered light passes through the second filter 14 to the second signal detection unit 15, an optical image (speckle image) is generated and transmitted to the PC, and then the optical image (speckle image) is processed into a digital image (speckle image) by the PC.
The multiphoton blood flow microscopic imaging system provided by the embodiment of the utility model is a multimode microscopic imaging system, and can observe the blood flow change of an irradiated area in real time through a speckle image, and simultaneously, the three-dimensional spatial distribution of blood vessels of the irradiated area is mapped through a three-dimensional image.
In the embodiment of the utility model, the multiphoton blood flow microscopic imaging system further comprises a laser processing unit 7 for collimating laser light, a coaxially arranged scanning field lens 8 and a barrel lens 9.
The laser processing unit 7 is arranged between the first laser 1 and the two-dimensional scanning unit 2 and comprises a laser collimation system 71, an adjustable laser attenuator 72, a spectroscope 73 and a laser power meter 74 electrically connected with the control unit 6. The laser emitted by the first laser 1 is collimated by a laser collimation system 71 to obtain a collimated laser beam, and then the laser beam is adjusted by an adjustable laser attenuator 72 to have proper optical power, and the laser beam is output to the two-dimensional scanning unit 2 after passing through a spectroscope 73. The collimated laser light passes through the beam splitter 73 and also a small portion of the collimated laser light enters the laser power meter 74 to monitor the laser power in real time. The split ratio of the beam splitter 73 may be 90:10 or 99:1, etc.
The optical axes of the scan field lens 8 and the barrel lens 9 coincide with the optical axis of the zoom objective lens 4, and are disposed between the two-dimensional scanning unit 2 and the first dichroic mirror 3. The scan field lens 8 has the function of eliminating scan distortion and flattening. The center of the two-dimensional scanning unit 2 is arranged at the focus on one side of the scanning field lens 8, the reflection angle of the first vibrating mirror 21 and the reflection angle of the second vibrating mirror 22 are changed, and the incident angle of the laser beam entering the scanning field lens 8 can be changed, so that two-dimensional scanning of a sample is realized. The cylindrical lens 9 has the functions of transition and amplification, the focus of the cylindrical lens 9 coincides with the focus of the other side of the scanning field lens 8, and the laser beam is emitted from the cylindrical lens 9 in a parallel light mode, passes through the first dichroic mirror 3 and enters the zoom objective lens 4.
Further, the second dichroic mirror 16 and the polarizing beam splitter 13 are disposed in a space between the scan field lens 8 and the barrel lens 9. Further, the multiphoton blood flow microscopic imaging system further includes a mirror 10. A mirror 10 is provided between the two-dimensional scanning unit 2 and the scan field lens 8 to change the direction of the laser beam according to the space requirement.
According to the embodiment of the utility model, the speckle image of blood flow is obtained through laser speckle blood flow imaging, the three-dimensional image of blood vessel distribution is obtained through multiphoton microscopic imaging, and the speckle image of blood flow and the three-dimensional image of blood vessel distribution are synthesized, so that the observation and research on tiny blood vessels are more accurate and visual.
While embodiments of the present utility model have been shown and described above, it will be understood that the above embodiments are illustrative and not to be construed as limiting the utility model, and that variations, modifications, alternatives and variations may be made to the above embodiments by one of ordinary skill in the art within the scope of the utility model.
Claims (10)
1. A multiphoton blood flow microscopy imaging system, comprising: the device comprises a first laser (1), a two-dimensional scanning unit (2), a first dichroic mirror (3), a zoom objective lens (4), a first signal detection unit (5), a second laser (11), a second dichroic mirror (16), a polarization spectroscope (13), a second signal detection unit (15) and a control unit (6);
the first laser (1), the two-dimensional scanning unit (2), the zoom objective lens (4), the first signal detection unit (5), the second laser (11) and the second signal detection unit (15) are respectively and electrically connected with the control unit (6);
the two-dimensional scanning unit (2) is arranged at one side of the first dichroic mirror (3) away from the zoom objective lens (4);
the optical axis of the first signal detection unit (5) is perpendicular to the optical axis of the zoom objective lens (4) and intersects the first dichroic mirror (3);
the optical axis of the second laser (11) is perpendicular to the optical axis of the zoom objective (4) and intersects the second dichroic mirror (16);
the optical axis of the second signal detection unit (15) is perpendicular to the optical axis of the zoom objective lens (4) and is intersected with the polarization beam splitter (13);
after passing through the two-dimensional scanning unit (2), excitation light emitted by the first laser (1) irradiates a sample through the first dichroic mirror (3) and the zoom objective lens (4), and fluorescence emitted by the excited sample transmits through the zoom objective lens (4) and is reflected at the first dichroic mirror (3) to enter the first signal detection unit (5);
after being reflected by the second dichroic mirror (16), the laser light emitted by the second laser (11) irradiates the sample through the zoom objective lens (4) to be scattered, and the scattered light passes through the zoom objective lens (4) and is reflected at the polarization beam splitter (13) to enter the second signal detection unit (15).
2. The multiphoton blood flow microscopic imaging system according to claim 1, wherein the first dichroic mirror (3), the second dichroic mirror (16) and the polarizing beam splitter (13) are arranged along an optical axis of the zoom objective lens (4), and the second dichroic mirror (16) is arranged at a side of the polarizing beam splitter (13) facing away from the first dichroic mirror (3).
3. The multiphoton blood flow microscopic imaging system according to claim 2, wherein the second dichroic mirror (16) and the polarizing beam splitter (13) are arranged at a side of the first dichroic mirror (3) facing away from the zoom objective (4).
4. The multiphoton blood flow microscopic imaging system according to claim 1, wherein the two-dimensional scanning unit (2) includes a first galvanometer (21) and a second galvanometer (22) electrically connected with the control unit (6), respectively; the first vibrating mirror (21) and the second vibrating mirror (22) are placed in an orthogonal mode, the reflection angle of the first vibrating mirror (21) is changed to conduct X-axis direction scanning, and the reflection angle of the second vibrating mirror (22) is changed to conduct Y-axis direction scanning.
5. The multiphoton blood flow microscopic imaging system according to claim 1, wherein the first signal detection unit (5) includes a converging lens (51) and a photomultiplier tube (52) coaxially disposed.
6. The multiphoton blood flow microscopic imaging system according to claim 5, wherein the first signal detection unit (5) further includes a first filter (53) coaxially disposed with the converging lens (51); the first optical filter (53) is arranged on one side of the converging lens (51) away from the photomultiplier (52).
7. The multiphoton blood flow microscopic imaging system according to claim 1, further comprising a laser processing unit (7) for collimating laser light, the laser processing unit (7) being arranged between the first laser (1) and the two-dimensional scanning unit (2).
8. The multiphoton blood flow microscopic imaging system according to claim 7, wherein the laser processing unit (7) includes a laser collimation system (71), an adjustable laser attenuator (72), a beam splitter (73), and a laser power meter (74).
9. The multiphoton blood flow microscopic imaging system according to claim 1, further comprising a scan field lens (8) and a barrel lens (9) coaxially arranged; the optical axes of the scanning field lens (8) and the barrel lens (9) are coincident with the optical axis of the zoom objective lens (4), and are arranged between the two-dimensional scanning unit (2) and the first dichroic mirror (3).
10. The multiphoton blood flow microscopic imaging system according to claim 9, further comprising a mirror (10), the mirror (10) being disposed between the two-dimensional scanning unit (2) and the scan field lens (8).
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| Application Number | Priority Date | Filing Date | Title |
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| CN202321685828.6U CN220572188U (en) | 2023-06-30 | 2023-06-30 | Multiphoton blood flow microscopic imaging system |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202321685828.6U CN220572188U (en) | 2023-06-30 | 2023-06-30 | Multiphoton blood flow microscopic imaging system |
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| CN220572188U true CN220572188U (en) | 2024-03-12 |
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| CN202321685828.6U Active CN220572188U (en) | 2023-06-30 | 2023-06-30 | Multiphoton blood flow microscopic imaging system |
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