CN115165826A - Near-infrared multichannel synchronous microscopic imaging system - Google Patents
Near-infrared multichannel synchronous microscopic imaging system Download PDFInfo
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- CN115165826A CN115165826A CN202210746964.5A CN202210746964A CN115165826A CN 115165826 A CN115165826 A CN 115165826A CN 202210746964 A CN202210746964 A CN 202210746964A CN 115165826 A CN115165826 A CN 115165826A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N21/6456—Spatial resolved fluorescence measurements; Imaging
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- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/0004—Microscopes specially adapted for specific applications
- G02B21/002—Scanning microscopes
- G02B21/0024—Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
- G02B21/0052—Optical details of the image generation
- G02B21/0076—Optical details of the image generation arrangements using fluorescence or luminescence
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N2021/6463—Optics
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N2021/6463—Optics
- G01N2021/6478—Special lenses
Abstract
The invention relates to a near-infrared multichannel synchronous microscopic imaging system, which relates to the technical field of microscopic optical imaging and comprises the following components: the device comprises a light-shading structure, a multi-channel imaging unit, a lens barrel lens, a switching type dichroic mirror filter unit, a switchable objective lens unit, a multi-channel excitation unit and an operation object translation stage, wherein the multi-channel imaging unit, the lens barrel lens, the switching type dichroic mirror filter unit, the switchable objective lens unit, the multi-channel excitation unit and the operation object translation stage are arranged in the light-shading structure; the multi-channel imaging unit includes: the camera set and the first dichroic mirror set; the multi-channel excitation unit comprises: an optical waveguide lens group and a second dichroic lens group. The imaging system can realize multi-component simultaneous and same-position imaging and detection.
Description
Technical Field
The invention relates to the technical field of microscopic optical imaging, in particular to a near-infrared multichannel synchronous microscopic imaging system.
Background
In the field of life science, optical imaging has the advantages of real-time feedback, simplicity in operation, non-invasion and the like. Early optical imaging used fluorescent probes emitting at 400nm to 900nm, but the biological tissue had high absorption and scattering of photons in this wavelength band, resulting in low imaging resolution and penetration depth. In recent years, scientists have found that photons of 1000nm to 1700nm have low scattering, absorption and auto-fluorescence in biological tissues, and fluorescent materials based on the emission of the wave band can realize optical imaging of deep tissues, high resolution and high signal-to-noise ratio. At present, wide-field microscopic imaging is developed more completely, and high-resolution and dynamic real-time observation of living biological samples can be realized. However, the existing near-infrared wide-field microscope system in the market can only realize the single excitation/single fluorescence imaging function, but cannot realize the imaging and detection of multiple components at the same time and the same position.
Disclosure of Invention
The invention aims to provide a near-infrared multichannel synchronous microscopic imaging system which can realize simultaneous imaging and detection of multiple components at the same time and position.
In order to achieve the purpose, the invention provides the following scheme:
a near-infrared multichannel synchronized microscopy imaging system, the imaging system comprising:
the device comprises a light-shading structure, a multi-channel imaging unit, a lens barrel lens, a switching type dichroic mirror filter unit, a switchable objective lens unit, a multi-channel excitation unit and an operation object translation stage, wherein the multi-channel imaging unit, the lens barrel lens, the switching type dichroic mirror filter unit, the switchable objective lens unit, the multi-channel excitation unit and the operation object translation stage are arranged in the light-shading structure;
the multi-channel imaging unit includes: the camera set and the first dichroic mirror set;
the multi-channel excitation unit includes: an optical waveguide lens group and a second dichroic lens group;
the optical waveguide lens group is used for adjusting the divergence angle and the light source diameter of the exciting light, the exciting light reaches the switching type dichroic mirror filter unit after several beams of exciting light are combined through the second dichroic mirror, the combined beams are reflected through the third dichroic mirror in the switching type dichroic mirror filter unit, and are converged to the plane of the operation objective translation stage through the objective lens in the switchable objective lens unit, the exciting light is aligned to the position to be measured through the operation objective translation stage under the adjustment of XY biaxial movement, the focusing is completed by adjusting the Z axis, the fluorescent light generated by the exciting fluorescent probe at the imaging position is collimated into parallel light through the switchable objective lens unit, the non-fluorescent interference is filtered through the dichroic mirror in the switching type dichroic mirror unit and the filter, and the parallel light is converged to the focal plane of the camera set through the lens barrel lens, and the imaging process is completed.
Optionally, the light-shielding structure is a box body.
Optionally, the camera comprises a plurality of near infrared cameras.
Optionally, the first dichroic mirror group includes a plurality of first dichroic mirrors, and the second dichroic mirror group includes a plurality of second dichroic mirrors.
Optionally, the manipulation stage comprises a three-axis combined motion including three dimensions of X-direction, Y-direction and Z-direction.
Optionally, the switchable objective unit is a 10 or 25 or 50 times near-infrared objective.
Optionally, the switching dichroic mirror optical filter unit includes a third dichroic mirror and a long-pass optical filter.
Optionally, the cut-off wavelength of the third dichroic mirror is 1000nm to 1500nm, and the cut-off wavelength of the optical filter is 1000nm to 1500nm.
According to the specific embodiment provided by the invention, the invention discloses the following technical effects:
the invention discloses a near-infrared multichannel synchronous microscopic imaging system, which comprises: the device comprises a light shading structure, a multi-channel imaging unit, a lens barrel lens, a switching type dichroic mirror filter unit, a switchable objective lens unit, a multi-channel excitation unit and an operation objective translation stage, wherein the multi-channel imaging unit, the lens barrel lens, the switching type dichroic mirror filter unit, the switchable objective lens unit, the multi-channel excitation unit and the operation objective translation stage are arranged in the light shading structure; the multi-channel imaging unit includes: the camera set and the first dichroic mirror set; the multi-channel excitation unit includes: an optical waveguide lens group and a second dichroic lens group; the optical waveguide lens group is used for adjusting the divergence angle and the light source diameter of exciting light, a plurality of beams of exciting light reach the switching type dichroic mirror filter unit after being combined by the second dichroic mirror, and then are reflected by the third dichroic mirror in the switching type dichroic mirror filter unit, and are converged to the plane of the operation objective translation stage by the objective lens in the switchable type objective lens unit, the exciting light is aligned to the position to be measured by the operation objective translation stage under the adjustment of XY two-axis motion, the focusing is completed by adjusting the Z axis, the fluorescent light generated by the fluorescent probe at the imaging position is collimated into parallel light by the switchable type objective lens unit, the non-fluorescent interference is filtered by the dichroic mirror in the switching type dichroic mirror filter unit, and is converged to the focal plane of the camera unit by the lens cone lens, the imaging process is completed, and the imaging and the detection of multiple components at the same time and the same position can be realized.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art that other drawings can be obtained according to these drawings without creative efforts.
FIG. 1 is a schematic structural diagram of a near-infrared multichannel synchronous microscopic imaging system according to an embodiment of the present invention;
FIG. 2 is a schematic diagram of synchronous two-channel emission imaging of elastic fiber and blood of living capillary vessel based on a near-infrared multichannel synchronous microscopic imaging system according to an embodiment of the present invention;
fig. 3 is a schematic diagram of synchronous dual-channel excitation imaging of elastic fiber and blood of living capillary vessels based on a near-infrared multichannel synchronous microscopic imaging system in the embodiment of the invention.
Description of the symbols:
the device comprises a near-infrared camera-1, a first dichroic mirror-2, a tube lens-3, a switching dichroic mirror filter unit-4, a switching objective lens unit-5, a second dichroic mirror-6, an optical waveguide lens group-7, an operation objective translation stage-8 and a light shading structure 9.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without making any creative effort based on the embodiments in the present invention, belong to the protection scope of the present invention.
The invention aims to provide a near-infrared multichannel synchronous microscopic imaging system which can realize simultaneous imaging and detection of multiple components at the same time and the same position.
In order to make the aforementioned objects, features and advantages of the present invention more comprehensible, the present invention is described in detail with reference to the accompanying drawings and the detailed description thereof.
Fig. 1 is a schematic structural diagram of a near-infrared multichannel synchronous microscopic imaging system according to an embodiment of the present invention, and as shown in fig. 1, the system includes:
the device comprises a light-shading structure 9, and a multi-channel imaging unit, a tube lens, a switching dichroic mirror filter unit, a switchable objective lens unit, a multi-channel excitation unit and an operation object stage translation which are arranged in the light-shading structure 9;
the multi-channel imaging unit includes: the camera set and the first dichroic mirror set;
the multi-channel excitation unit comprises: an optical waveguide lens group and a second dichroic lens group;
the optical waveguide lens group is used for adjusting the divergence angle and the light source diameter of the exciting light, the exciting light reaches the switching type dichroic mirror filter unit after several beams of exciting light are combined through the second dichroic mirror, the combined beams are reflected through the third dichroic mirror in the switching type dichroic mirror filter unit, and are converged to the plane of the operation objective translation stage through the objective lens in the switchable objective lens unit, the exciting light is aligned to the position to be measured through the operation objective translation stage under the adjustment of XY biaxial movement, the focusing is completed by adjusting the Z axis, the fluorescent light generated by the exciting fluorescent probe at the imaging position is collimated into parallel light through the switchable objective lens unit, the non-fluorescent interference is filtered through the dichroic mirror in the switching type dichroic mirror unit and the filter, and the parallel light is converged to the focal plane of the camera set through the lens barrel lens, and the imaging process is completed.
Wherein, light-resistant structure 9 is the box, and light-resistant structure 9 covers whole part, avoids the interference of ambient light to imaging quality.
In the imaging process, a second dichroic mirror group is arranged in front of the optical waveguide lens group 7, and at least three channels of exciting light can be switched by a second dichroic mirror 6 in the second dichroic mirror group, so that multichannel time sequence control excitation is realized. Similarly, a first dichroic mirror group is arranged in front of the near-infrared imaging camera 1, and synchronous fluorescence imaging of at least 3 different wave bands at the same time and the same position can be completed through a first dichroic mirror 2 in the first dichroic mirror group.
The switching dichroic mirror filter unit 4 can preset at most 8 groups of dichroic mirrors with different wavelengths and filter combinations according to the excitation/emission properties of the fluorescent probe.
The near-infrared multichannel synchronous microscopic imaging system comprises bright field and dark field observation shot by a near-infrared imaging camera, wherein the bright field is a near-infrared laser used as an illumination light source, and a sample is observed through scattering and reflected light; and dark field is that after a fluorescent probe and a corresponding filter are combined, imaging analysis is carried out.
The near-infrared multichannel synchronous microscopic imaging system comprises a switchable objective unit 5 with at most 6 channels, and objective lenses with different magnification factors can be switched according to requirements.
The near-infrared multichannel synchronous microscopic imaging system comprises three-axis combined motion comprising three dimensions of X direction, Y direction and Z direction, and Z-axis motion has two adjusting modes of coarse adjustment and fine adjustment, so that accurate imaging of different parts of a living body can be realized.
The camera includes a plurality of near-infrared cameras 1, and the number of the near-infrared cameras in this embodiment is 3.
The first dichroic mirror group comprises a plurality of first dichroic mirrors, and the second dichroic mirror group comprises a plurality of second dichroic mirrors. In this embodiment, the number of the first dichroic mirrors is 2, and the number of the second dichroic mirrors is also 2.
The near-infrared multichannel synchronous microscopic imaging system formed by the device has the following operation modes of multichannel emission imaging:
the used fluorescent probes are Er-doped nanoparticles and Ho-doped nanoparticles, the excitation wavelength is 980nm, and the emission wavelengths are 1525nm and 1180nm respectively.
mPEG is modified on the surface of the fluorescent probe to realize water phase transfer.
The Er probe is used for marking the artery elastic fiber, and the Ho probe is dispersed in blood.
The laser in the multi-channel excitation unit is 980nm laser.
The switching dichroic mirror optical filter unit 4 adopts an 1100nm long-pass dichroic mirror, and an 1100nm long-pass optical filter is arranged above the switching dichroic mirror optical filter unit.
The switchable objective unit 5 is a 10-fold near-infrared objective.
The first dichroic mirror 2 in the multi-channel imaging unit selects a long-pass dichroic mirror with the cut-off wavelength of 1300 nm.
A near-infrared camera 1 in the multi-channel imaging unit adopts an InGaAs camera, er probe fluorescence is detected above a first dichroic mirror, and Ho probe fluorescence is detected by the other camera.
And placing the processed observed sample on an operation translation table 8, opening the instrument, starting a laser light source, completing preliminary focusing through the z-axis translation of the operation translation table 8, and searching an imaging position through the x-axis and y-axis movement.
After the initial focusing is completed, the laser power density is adjusted, the image shot by the near-infrared imaging camera 1 is observed through computer software, and the precise focusing is performed to obtain clear fluorescent imaging, and the result is shown in fig. 2.
The multi-channel excitation imaging operation mode comprises the following steps:
the fluorescent probe is composed of two Er-doped nanoparticles (1) and (2), the excitation wavelength of the probe (1) is 808nm, the excitation wavelength of the probe (2) is 980nm, and the emission wavelengths of the probe (1) and the probe (2) are 1525nm.
mPEG is modified on the surfaces of the probe (1) and the probe (2) to realize water transfer.
The 808nm excitation probe (1) is used for labeling the artery elastic fiber, and the 980nm excitation probe (2) is dispersed in blood.
The lasers in the multi-channel excitation unit are 980nm and 808nm lasers, and the second dichroic mirror 6 is a long-pass dichroic mirror with the cut-off wavelength of 900 nm.
The switching dichroic mirror filter unit 4 adopts an 1100nm long-pass dichroic mirror, and an 1100nm long-pass filter is arranged above the switching dichroic mirror filter unit.
The switchable objective unit 5 is a 25-fold near-infrared objective.
The near-infrared camera 1 in the multi-channel imaging unit adopts an InGaAs camera.
And placing the processed observed sample on an operation translation table 8, starting the instrument, respectively starting a laser light source, completing preliminary focusing through the z-axis translation of the operation translation table 8, and searching an imaging position through the x-axis and y-axis movement.
After the initial focusing is completed, the laser power density is adjusted, the image shot by the near-infrared imaging camera 1 is observed through computer software, and the precise focusing is performed to obtain clear fluorescent image, and the result is shown in fig. 3.
In the present specification, the embodiments are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other.
The principle and the embodiment of the present invention are explained by applying specific examples, and the above description of the embodiments is only used to help understanding the method and the core idea of the present invention; meanwhile, for a person skilled in the art, according to the idea of the present invention, the specific embodiments and the application range may be changed. In view of the above, the present disclosure should not be construed as limiting the invention.
Claims (8)
1. A near-infrared multichannel synchronized microscopy imaging system, the imaging system comprising:
the device comprises a light-shading structure, a multi-channel imaging unit, a lens barrel lens, a switching type dichroic mirror filter unit, a switchable objective lens unit, a multi-channel excitation unit and an operation object translation stage, wherein the multi-channel imaging unit, the lens barrel lens, the switching type dichroic mirror filter unit, the switchable objective lens unit, the multi-channel excitation unit and the operation object translation stage are arranged in the light-shading structure;
the multi-channel imaging unit includes: the camera set and the first dichroic mirror set;
the multi-channel excitation unit includes: an optical waveguide lens group and a second dichroic lens group;
the optical waveguide lens group is used for adjusting the divergence angle and the light source diameter of exciting light, a plurality of beams of exciting light reach the switching type dichroic mirror filter unit after being combined by the second dichroic mirror, and then are reflected by the third dichroic mirror in the switching type dichroic mirror filter unit, and are converged to the plane of the operation objective translation stage by the objective lens in the switchable type objective lens unit, the exciting light is aligned to the position to be measured by the operation objective translation stage under the adjustment of XY two-axis motion, the focusing is completed by adjusting the Z axis, the fluorescent light generated by the fluorescent probe at the imaging position is collimated into parallel light by the switchable type objective lens unit, and is transmitted by the dichroic mirror in the switching type dichroic filter unit and filtered by the optical filter to remove non-fluorescent interference, and is converged to the focal plane of the camera unit by the lens cone lens, and the imaging process is completed.
2. The near-infrared multichannel synchronous microscopic imaging system according to claim 1, characterized in that the light-shielding structure is a box body.
3. The near-infrared multichannel synchronized microscopy imaging system of claim 1, wherein the camera comprises a plurality of near-infrared cameras.
4. The near-infrared multichannel synchronized microscopy imaging system of claim 1, wherein the first set of dichroic mirrors comprises a plurality of first dichroic mirrors and the second set of dichroic mirrors comprises a plurality of second dichroic mirrors.
5. The near-infrared multichannel synchronized microscopy imaging system of claim 1, wherein the manipulation stage translation stage comprises a three-axis compound motion comprising three dimensions of X, Y, and Z.
6. The near-infrared multichannel synchronized microscopy imaging system according to claim 1, wherein the switchable objective unit is a 10 or 25 or 50 times near-infrared objective.
7. The near-infrared multichannel synchronized microscopy imaging system of claim 1, wherein the switched dichroic mirror filter unit comprises a third dichroic mirror and a long pass filter.
8. The near-infrared multichannel synchronous microscopic imaging system according to claim 7, wherein the third dichroic mirror has a cutoff wavelength of 1000nm to 1500nm, and the filter has a cutoff wavelength of 1000nm to 1500nm.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN117348224A (en) * | 2023-12-04 | 2024-01-05 | 成都丹诺迪医疗科技有限公司 | Microscopic device |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN117348224A (en) * | 2023-12-04 | 2024-01-05 | 成都丹诺迪医疗科技有限公司 | Microscopic device |
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