CN114911044A - Staring type fast light sheet confocal high-dimensional imaging system - Google Patents

Staring type fast light sheet confocal high-dimensional imaging system Download PDF

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Publication number
CN114911044A
CN114911044A CN202210368281.0A CN202210368281A CN114911044A CN 114911044 A CN114911044 A CN 114911044A CN 202210368281 A CN202210368281 A CN 202210368281A CN 114911044 A CN114911044 A CN 114911044A
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lens
light
scanning
unit
reflector
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何赛灵
罗晶
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Zhejiang University ZJU
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Zhejiang University ZJU
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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
    • G02B21/0032—Optical details of illumination, e.g. light-sources, pinholes, beam splitters, slits, fibers
    • 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
    • G02B21/0036—Scanning details, e.g. scanning stages
    • 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
    • G02B21/0036—Scanning details, e.g. scanning stages
    • G02B21/0048—Scanning details, e.g. scanning stages scanning mirrors, e.g. rotating or galvanomirrors, MEMS mirrors
    • 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
    • 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
    • G02B21/008—Details of detection or image processing, including general computer control
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/24—Base structure
    • G02B21/26—Stages; Adjusting means therefor
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements
    • G02B21/365—Control or image processing arrangements for digital or video microscopes

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  • Physics & Mathematics (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Optics & Photonics (AREA)
  • Engineering & Computer Science (AREA)
  • Computer Vision & Pattern Recognition (AREA)
  • Multimedia (AREA)
  • General Engineering & Computer Science (AREA)
  • Microscoopes, Condenser (AREA)

Abstract

The invention discloses a staring type fast light sheet confocal high-dimensional imaging system which sequentially comprises an excitation unit, a rear end receiving unit, a middle scanning unit and a front optical unit; the excitation unit sequentially comprises a collimating lens, a first cylindrical lens, a second cylindrical lens and a first reflector; the middle scanning unit sequentially comprises a fourth cylindrical lens, a galvanometer system, a scanning lens and a telecentric lens, wherein the scanning lens and the telecentric lens are used for optimizing light beams and expanding beams, and the galvanometer system is used for one-dimensional scanning; the preposed optical unit comprises an object stage, a microobjective and a second reflector; the rear end receiving unit sequentially comprises a dichroic mirror, an optical filter, a third cylindrical lens and a hyperspectral light splitting module. The invention overcomes the problems of low scanning speed and single data of the traditional point confocal system, has high spectral resolution and high depth resolution, and has great application value in the measurement fields of in-situ microorganism detection, industrial sample microstructure detection and the like.

Description

Staring type fast light sheet confocal high-dimensional imaging system
Technical Field
The invention belongs to the technical field of optics, and particularly relates to a staring type fast optical sheet confocal high-dimensional imaging system.
Background
The confocal laser scanning microscope is a novel instrument with high photosensitivity and high resolution developed in nearly more than ten years. The laser is used as a light source and consists of a confocal imaging scanning system, an electronic optical system and a microcomputer image analysis system. The light beam is focused and then falls on a tiny point of a sample (a tissue slab or a cell) with different depths, and the movement scanning is carried out, the image formed by the reflected light of any point in the sample can be accurately received and generate a signal through electric signal color imaging, and the image signal is obtained through processing. Confocal microscope systems, however, are slow to acquire, inefficient, and lack means for analyzing samples. At present, hyperspectral imaging is based on a multichannel spectral technology, optical imaging and spectral measurement are integrated, and image information and corresponding spectral information of a target can be acquired simultaneously. The hyperspectral imaging can analyze, measure and process the structure and the components of a substance, has the advantages of high analysis precision, wide measurement range and the like, and is widely applied to the fields of petroleum, materials, agriculture, geological exploration, biochemistry, medical sanitation, environmental protection, safety detection and the like.
Disclosure of Invention
In order to overcome the defects in the prior art, the invention discloses a gaze type fast optical sheet confocal high-dimensional imaging system.
A staring type fast light sheet confocal high-dimensional imaging system sequentially comprises an excitation unit, a rear end receiving unit, a middle scanning unit and a front optical unit; the excitation unit sequentially comprises a collimating lens, a first cylindrical lens, a second cylindrical lens and a first reflector; the middle scanning unit sequentially comprises a fourth cylindrical lens, a vibrating mirror system, a scanning lens and a telecentric lens, the scanning lens and the telecentric lens are used for optimizing light beams and expanding beams, and the vibrating mirror system is used for one-dimensional scanning; the preposed optical unit comprises an objective table, a microscope objective and a second reflector, wherein the line excitation light emitted from the middle scanning unit is incident to the microscope objective after passing through the second reflector and is focused on a sample to be measured on the objective table to form a light sheet; the rear end receiving unit sequentially comprises a dichroic mirror, an optical filter, a third cylindrical lens and a hyperspectral light splitting module.
A staring type fast light sheet confocal high-dimensional imaging system, line excitation light is focused on the surface of a sample through a microscope objective to form a light sheet, a fluorescence signal or a reflected light signal at the focal plane of the microscope objective is received by a hyperspectral light-splitting module, light at other positions of the focal plane is shielded by a slit in the hyperspectral light-splitting module, and chromatographic scanning can be realized through axial displacement of the microscope objective.
The hyperspectral light splitting module sequentially comprises a slit, a collimating lens, a prism-grating-prism, a focusing lens and a camera, light is emitted from the middle scanning unit to the slit position and collimated into parallel light by the collimating lens, and after passing through the prism-grating-prism, the light with different wavelengths is focused on different positions of a camera photosurface by the focusing lens, so that a spectral image is formed.
The front-mounted optical unit comprises an object stage, a microscope objective, an electric displacement stage and a second reflector, wherein the electric displacement stage is used for adjusting the distance of a sample to be measured to realize focusing.
The front optical unit comprises an object stage, a microscope objective, a liquid zoom lens and a second reflector, wherein the liquid zoom lens is used for realizing rapid zooming.
The invention has the beneficial effects that:
the invention adopts the line confocal light sheet to scan the fluorescence or reflection map information of a sample to be detected, and combines the chromatography technology to carry out depth scanning, so as to fuse the fluorescence or reflection map data of the sample to be detected into a high-dimensional data cube. The system has the characteristics of high depth resolution, high spectral resolution, excellent data fusion precision and the like, and the new spectral morphology data volume greatly improves the accuracy of microscopic detection and analysis of substances.
The invention overcomes the problems of low acquisition speed, low detection efficiency, single data and the like of the traditional confocal system, combines the linear confocal technology with the grating line push-broom spectral imaging technology, and can detect the fluorescence spectrum or reflection spectrum data of a sample to be detected with high precision. The system can perform chromatographic scanning on the sample through the axial movement of the microscope objective, has important significance for analyzing the microscopic properties of the sample to be detected, and has great application value in the measurement fields of in-situ microbial detection, industrial sample microstructure detection and the like.
Drawings
FIG. 1 is a schematic diagram of a four-dimensional detection system for a staring confocal micro-topography spectrum;
in the figure, a collimating lens 1, a first cylindrical lens 2, a second cylindrical lens 3, a first reflecting mirror 4, a dichroic mirror 5, a filter 6, a third cylindrical lens 7, a hyperspectral spectroscopy module 8, a slit 9, a collimating lens 10, a prism-grating-prism 11, a focusing lens 12, a camera 13, a fourth cylindrical lens 14, a galvanometer system 15, a scanning lens 16, a telecentric lens 17, a second reflecting mirror 18, an electric displacement table 19, a microscope objective 20, an objective table 21, an excitation unit 22, a rear-end receiving unit 23, a middle scanning unit 24 and a front-end optical unit 25 are arranged.
Detailed Description
The invention is further elucidated with reference to the drawing.
As shown in fig. 1, a gaze-fixation type fast optical sheet confocal high-dimensional imaging system sequentially comprises an excitation unit 22, a rear end receiving unit 23, a middle scanning unit 24, and a front optical unit 25.
The excitation unit 22 sequentially comprises a collimating lens 1, a first cylindrical lens 2, a second cylindrical lens 3 and a first reflector 4, exciting light of a sample to be detected is led in from the excitation unit, and the propagation direction of laser is upward; exciting light of a sample to be detected is collimated into parallel light through the collimating lens 1 after being emitted from the optical fiber, the parallel light is expanded in a single direction through the first cylindrical lens 2 and the second cylindrical lens 3, the expanded laser beam passes through the dichroic mirror 5 in the rear-end receiving unit 23, and then the exciting light with short wavelength is reflected to the middle scanning unit 24.
The middle scanning unit 24 sequentially comprises a fourth cylindrical lens 14, a galvanometer system 15, a scanning lens 16 and a telecentric lens 17, the scanning lens 16 and the telecentric lens 17 optimize light beams and expand the light beams, and the galvanometer system 15 performs one-dimensional scanning.
The front optical unit 25 includes an object stage 21, a microscope objective 20, an electric displacement stage 19 and a second reflector 18, wherein the linear excitation light emitted from the middle scanning unit 24 is incident to the microscope objective 20 after passing through the second reflector 18 and is focused on the sample to be measured on the object stage 21 to form an optical sheet, the electric displacement stage 19 is used for adjusting the distance of the sample to be measured to realize focusing, and the linear fluorescence signal or the linear reflection light signal excited by the sample to be measured returns to the rear end receiving unit 23 along the original optical path.
The electric displacement table 19 in the front optical unit 25 can be replaced by a liquid zoom lens for realizing quick zooming and improving the speed of the system tomography.
The rear-end receiving unit 23 sequentially comprises a dichroic mirror 5, an optical filter 6, a third cylindrical lens 7 and a hyperspectral light splitting module 8, linear fluorescence or linear reflection light enters the dichroic mirror 5 in the rear-end receiving unit through a middle scanning unit 24 along an original light path, is focused at the slit position of the hyperspectral light splitting module through the third cylindrical lens 7, and is unfolded into a fluorescence spectrum or a reflection map on a camera light-sensitive surface.
The line excitation light passes through the focus of microobjective 20 and forms the slide on the sample surface, and only fluorescence signal or the reflected light signal of 20 focal planes departments of microobjective can be received by high spectrum beam splitting module 8, and the light of the other positions of focal plane can be sheltered from by the slit in high spectrum beam splitting module 8, can realize the chromatography scanning through microobjective's axial displacement.
The system adopts optical sheet scanning, and has high scanning efficiency and high acquisition speed.
The hyperspectral light splitting module 8 sequentially comprises a slit 9, a collimating lens 10, a prism-grating-prism 11, a focusing lens 12 and a camera 13, light is emitted from the middle scanning unit 24 to the slit position and collimated into parallel light by the collimating lens 10, and after passing through the prism-grating-prism 11, light with different wavelengths is focused on different positions of a light sensing surface of the camera 13 through the focusing lens 12, so that a spectral image is formed.
The embodiments in the above description can be further combined or replaced, and the embodiments are only described as preferred examples of the present invention, and do not limit the concept and scope of the present invention, and various changes and modifications made to the technical solution of the present invention by those skilled in the art without departing from the design concept of the present invention belong to the protection scope of the present invention. The scope of the invention is given by the appended claims and any equivalents thereof.

Claims (5)

1. A staring type fast light sheet confocal high-dimensional imaging system is characterized by sequentially comprising an excitation unit (22), a rear end receiving unit (23), a middle scanning unit (24) and a front optical unit (25);
the excitation unit (22) sequentially comprises a collimating lens (1), a first cylindrical lens (2), a second cylindrical lens (3) and a first reflector (4);
the middle scanning unit (24) sequentially comprises a fourth cylindrical lens (14), a galvanometer system (15), a scanning lens (16) and a telecentric lens (17), the scanning lens (16) and the telecentric lens (17) are used for optimizing light beams and expanding beams, and the galvanometer system (15) is used for one-dimensional scanning;
the preposed optical unit (25) comprises an object stage (21), a microscope objective (20) and a second reflector (18), wherein the line excitation light emitted from the middle scanning unit (24) enters the microscope objective (20) after passing through the second reflector (18) and is focused on a sample to be detected on the object stage (21) to form a light sheet;
the rear receiving unit (23) sequentially comprises a dichroic mirror (5), a filter (6), a third cylindrical lens (7) and a hyperspectral light splitting module (8).
2. The gaze-type rapid light sheet confocal high-dimensional imaging system according to claim 1, wherein line excitation light is focused on the surface of a sample through the microscope objective (20) to form a light sheet, a fluorescence signal or a reflected light signal at the focal plane of the microscope objective (20) is received by the hyperspectral spectroscopy module (8), light at other positions of the focal plane is blocked by a slit in the hyperspectral spectroscopy module (8), and tomographic scanning can be realized through axial displacement of the microscope objective.
3. The gaze-type rapid light sheet confocal high-dimensional imaging system according to claim 1, wherein the hyperspectral light splitting module (8) sequentially comprises a slit (9), a collimating lens (10), a prism-grating-prism (11), a focusing lens (12) and a camera (13), light is emitted from the middle scanning unit (24) to the slit position and is collimated into parallel light by the collimating lens (10), and after passing through the prism-grating-prism (11), light with different wavelengths is focused on different positions of a light sensing surface of the camera (13) through the focusing lens (12), so that a spectral image is formed.
4. The gaze-type fast light-sheet confocal high-dimensional imaging system according to claim 1, wherein the front optical unit (25) comprises a stage (21), a microscope objective (20), an electric displacement stage (19) and a second reflector (18), and the electric displacement stage (19) is used for adjusting the distance of the sample to be measured to realize focusing.
5. The gaze-type fast light sheet confocal high-dimensional imaging system according to claim 1, wherein the front optical unit (25) comprises a stage (21), a microscope objective (20), a liquid zoom lens, and a second reflector (18), and the liquid zoom lens is used for realizing fast zooming.
CN202210368281.0A 2022-04-09 2022-04-09 Staring type fast light sheet confocal high-dimensional imaging system Pending CN114911044A (en)

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CN202210368281.0A CN114911044A (en) 2022-04-09 2022-04-09 Staring type fast light sheet confocal high-dimensional imaging system

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