CN106706589B - Fluorescence detection system for cell analyzer - Google Patents
Fluorescence detection system for cell analyzer Download PDFInfo
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- CN106706589B CN106706589B CN201710038022.0A CN201710038022A CN106706589B CN 106706589 B CN106706589 B CN 106706589B CN 201710038022 A CN201710038022 A CN 201710038022A CN 106706589 B CN106706589 B CN 106706589B
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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
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
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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
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
Abstract
The invention discloses a fluorescence detection system for a cell analyzer, which comprises an objective lens, an optical fiber, a concave mirror group, a cylindrical lens group, a spectroscope group, a band-pass filter group, a lens group, a detector group and the like. The invention separates the multi-wavelength fluorescence signals in space according to the wavelength and images the signals on the detector with 1 multiplied by the amplification factor, thus realizing the analysis and detection of fluorescence; the invention has the advantages of compact structure, small volume, simple debugging and the like; the number of the analysis and detection fluorescence wavelengths can be flexibly adjusted according to the actual needs of clients.
Description
Technical Field
The invention relates to the field of medical instruments, in particular to a fluorescence detection system for a cell analyzer.
Background
In the biological and medical fields, flow cytometers are often used to quantitatively count and analyze the types and numbers of biological cells. The flow cytometry adopts fluorescent reagent to dye the sample, the dyed sample particles pass through the detection area, and simultaneously, the detection area is irradiated by the laser beam, and different types of dyed sample particles emit fluorescent signals with different wavelengths, and the fluorescent signals with different wavelengths are mixed together and collected by the objective lens. Meanwhile, the spectroscope and the band-pass filter are adopted to divide fluorescence into fluorescence signals with different wavelengths, then the fluorescence signals with different wavelengths are analyzed one by one, and the types and the numbers of particles in a sample are counted rapidly through software calculation.
US patent 6683314 uses a star light path system to collect the detected fluorescent signal. In the invention, a long-focus objective lens is used for collecting fluorescence, but the problems of aberration and divergence angle of the fluorescence in the transmission process limit the number of fluorescence wavelengths which can be detected, so that the invention can only detect fluorescence with no more than 6 wavelengths; meanwhile, the fluorescent light spot incident on the photosensitive surface of the detector is larger than 3mm. Therefore, the technology provided by the invention can only adopt a photomultiplier tube (PMT) with large surface area as a detector, so that the whole system has larger volume.
CN 103091311A disclosed in chinese patent document designs a fluorescence collection detection system, which adopts a biprism group beam splitting method to sequentially arrange fluorescence in a wide spectral range according to wavelength in space. The same number of optical fiber groups are used to transmit fluorescent signals according to the number of fluorescent wavelengths to be detected. The whole system has higher requirements on the position accuracy of the biprism and the optical fiber group, so that the adjustment difficulty of the whole system is further increased.
Disclosure of Invention
The invention aims to provide a fluorescence detection system for a cell analyzer, which can realize that the number of detected fluorescence wavelengths is not limited by aberration and divergence angle in the transmission process, and meanwhile, the whole system has the advantages of compact structure, small volume, simple debugging and the like.
The aim of the invention is achieved by the following technology:
this kind of fluorescence detection system for cell analysis appearance, characterized by: the fluorescent detection system comprises an objective lens, an optical fiber, a concave lens group, a cylindrical lens group, a spectroscope group, a band-pass filter group, a lens group and a detector group which form a detection area, and the fluorescent detection system formed by the components collects and detects fluorescence emitted by dyed sample particles passing through the detection area after laser irradiation;
wherein:
the objective lens is used for collecting and collimating fluorescence generated after the sample particles are excited by the laser;
the optical fiber comprises an incident end face and an emergent end face, and is used for transmitting fluorescent light collimated by the objective lens;
the concave mirror group comprises a plurality of concave mirrors which are arranged in an array and are used for reflecting fluorescence, the concave surfaces of the concave mirrors are plated with total reflection films, and the concave surfaces of the concave mirrors face to the optical fibers;
the cylindrical lens group comprises a plurality of plano-convex cylindrical lenses which are arranged in an array and used for correcting astigmatism, and both surfaces of each cylindrical lens are plated with an antireflection film system;
the spectroscope group comprises a plurality of spectroscopes which are arranged in an array, each spectroscope is respectively coated with a film according to the fluorescent wavelength to be detected, so that fluorescent signals larger than a certain wavelength pass through, and fluorescent signals smaller than the wavelength are reflected;
the band-pass filter set comprises a plurality of band-pass filters which are arranged in an array, each filter is respectively coated according to the fluorescent wavelength to be detected, so that the fluorescent light with a specific wavelength passes, all the fluorescent light with other wavelengths are respectively coated with the fluorescent wavelength which is reversely detected, so that the fluorescent light with the specific wavelength passes, and all the fluorescent light with other wavelengths is reflected;
the lens group comprises a plurality of aspherical lenses which are arranged in an array, two surfaces of each aspherical lens are plated with an antireflection film system, and the aspherical lenses image fluorescent light spots on the spectroscope on a photosensitive surface of the detector;
the detector group comprises a plurality of detectors which are arranged in an array, the detectors are arranged corresponding to the fluorescent wavelengths divided by the band-pass filter, and the detectors are used for receiving fluorescent signals and converting the fluorescent signals into electric signals.
The spectroscope is arranged near the position of the double focal length of the concave mirror corresponding to the spectroscope.
The photosensitive surface of the detector is positioned near the position of the double focal length of the corresponding aspheric lens.
The spectroscope is positioned near the position of the double focal length of the corresponding aspheric lens.
The incident end face of the optical fiber is positioned near the focal point of the objective lens; the emergent end face of the optical fiber is positioned near the position of the double focal length of the concave mirror.
The objective lens has a high Numerical Aperture (NA) of 0.5-1.4.
The optical fiber has a relatively low numerical aperture and NA of 0.05-0.15.
The aspherical lenses arranged in an array have higher numerical aperture and NA of 0.5-0.9.
The convex curvature faces of the cylindrical lenses in the cylindrical lens array face away from the detector.
The photosensitive face of the detector array faces the lens; the detector is a photomultiplier tube (PMT) or an Avalanche Photodiode (APD).
According to the invention provided by the technical scheme, the fluorescence detection system adopts a Z-shaped light path, uses the low numerical aperture optical fiber to output a fluorescence signal, adopts the cylindrical lens to correct the divergence angle and astigmatism of fluorescence, and the aspheric lens corrects the chromatic aberration, spherical aberration and other aberration of the fluorescence signal, finally, the fluorescence signal is sequentially arranged in space according to the wavelength and imaged on the photosensitive surface of the detector with the 1x amplification rate, and the size of a fluorescence spot on the photosensitive surface of the detector is basically equal to the core diameter of the optical fiber. The invention can therefore use a small surface area detector (< 1mm, such as an avalanche diode APD) as the detector to greatly reduce the system volume; the invention can realize that the number of the detected fluorescence wavelengths can be increased or decreased according to the needs of clients, can realize the detection of fluorescence signals with one or more wavelengths at the same time, and has better client adaptability; meanwhile, the optical device has the advantages of low requirement on adjustment precision, better cost advantage, simple production process and the like.
Drawings
FIG. 1 is a schematic diagram of a combination of a flow cell, objective lens and optical fiber for a fluorescence detection system of a cell analyzer according to the present invention;
FIG. 2 is a schematic diagram showing the internal structure of a fluorescence detection system for a cell analyzer according to the present invention;
FIG. 3 is a schematic diagram of the optical path of the fluorescence signal transmitted in the system according to the present invention;
wherein 1A is a flow chamber; 1B is an objective lens; 2 is an optical fiber, 21 is an optical fiber incident end face, and 22 is an optical fiber emergent end face;
3 is a concave mirror group comprising concave mirrors 31, 32, 33, 34 and 35;
4 is a cylindrical lens group including cylindrical lenses 41, 42, 43, 44, and 45;
5 is a spectroscope group including spectroscopes 51, 52, 53, 54 and 55;
6 is a bandpass filter set including bandpass filters 61, 62, 63, 64, and 65;
7 is a lens group including aspherical lenses 71, 72, 73, 74 and 75;
8 are detector sets including detectors 81, 82, 83, 84 and 85.
Detailed Description
The invention will be described in further detail below with reference to the drawings by means of specific embodiments.
This kind of fluorescence detection system for cell analysis appearance, characterized by: the fluorescence detection system comprises an objective lens 1B, an optical fiber 2, a concave mirror group 3, a cylindrical lens group 4, a spectroscope group 5, a band-pass filter group 6, a lens group 7 and a detector group 8 which form a detection area, and the fluorescence detection system formed by the components collects and detects fluorescence emitted by dyed sample particles passing through the detection area after laser irradiation;
wherein:
the objective lens is used for collecting and collimating fluorescence generated after the sample particles are excited by the laser;
the optical fiber comprises an incident end face and an emergent end face, and is used for transmitting fluorescent light collimated by the objective lens;
the concave mirror group comprises a plurality of concave mirrors which are arranged in an array and used for reflecting fluorescent signals, wherein the concave surfaces of the concave mirrors are plated with total reflection films, and the concave surfaces of the concave mirrors face to the optical fibers;
the cylindrical lens group comprises a plurality of plano-convex cylindrical lenses which are arranged in an array and used for correcting astigmatism, and both surfaces of each cylindrical lens are plated with an antireflection film system;
the spectroscope group comprises a plurality of spectroscopes which are arranged in an array, each spectroscope is respectively coated with a film according to the fluorescent wavelength to be detected, so that fluorescent signals larger than a certain wavelength pass through, and fluorescent signals smaller than the wavelength reflect;
the band-pass filter set comprises a plurality of band-pass filters which are arranged in an array, each filter is respectively coated with a film according to the wavelength of fluorescence to be detected, so that fluorescence with a specific wavelength passes through, and fluorescence with other wavelengths is totally reflected;
the lens group comprises a plurality of aspherical lenses which are arranged in an array, wherein both surfaces of each aspherical lens are plated with an antireflection film system, and the aspherical lenses image fluorescent light spots on a spectroscope on a photosensitive surface of the detector;
the detector comprises a plurality of detectors which are arranged in an array, the detectors are arranged corresponding to the fluorescent wavelengths divided by the band-pass filter, and the detectors are used for receiving fluorescent signals and converting the fluorescent signals into electric signals.
As shown in fig. 1 and 2, a fluorescence detection system for a cell analyzer according to an embodiment of the present invention includes:
in the optical path are placed a flow cell 1A, an objective lens 1B, an optical fiber 2 (including an entrance end face 21 and an exit end face 22), a concave mirror group 3 (including 5 concave mirrors, respectively 31, 32, 33, 34, and 35), a cylindrical lens group 4 (including 5 cylindrical lenses, respectively 41, 42, 43, 44, and 45), a spectroscopic group 5 (including five spectroscopes, respectively 51, 52, 53, 54, and 55), a band-pass filter group 6 (including 5 band-pass filters, respectively 61, 62, 63, 64, and 65), a lens group 7 (including 5 aspherical lenses, respectively 71, 72, 73, 74, and 75), and a detector group 8 (including 5 detectors, respectively 81, 82, 83, 84, and 85).
Wherein, the concave surfaces of the concave mirrors are plated with a total reflection film system for reflecting fluorescent signals; the cylindrical lens and the aspheric lens are plated with an antireflection film system; the spectroscope is respectively coated with a film according to the fluorescent wavelength to be detected, so that fluorescent light with a wavelength larger than a certain wavelength passes through, and fluorescent signals with a wavelength smaller than the certain wavelength are reflected; the bandpass filters are respectively coated according to the fluorescent wavelength to be detected, so that the fluorescent light with a specific wavelength passes through, and the fluorescent light with other wavelengths is totally reflected;
the sample particles are dyed by adopting fluorescent reagent, the dyed sample particles pass through the flow chamber 1A, and simultaneously the flow chamber is irradiated by laser, and the sample particles emit fluorescent signals after being excited, wherein the fluorescent signals comprise broadband fluorescence with various wavelengths, and the wavelengths are lambda respectively 1 、λ 2 、λ 3 、λ 4 、λ 5 And lambda is 1 >λ 2 >λ 3 >λ 4 >λ 5 The method comprises the steps of carrying out a first treatment on the surface of the Collecting fluorescent signals generated by the sample particles by the objective lens 1B having a high numerical aperture, and collimating the fluorescent signals; the collimated fluorescence enters the optical fiber 2 through the optical fiber incident end face 21 and is output from the optical fiber emergent end face 22; the optical fiber incident end face 21 is located near the focal position of the objective lens, and the optical fiber exit end face 22 is located near the double focal position of the concave mirror.
The fluorescent signal (including the wavelength lambda) outputted from the optical fiber exit end face 22 1 、λ 2 、λ 3 、λ 4 、λ 5 ) At a certain angle to the concave mirror31 and is reflected; wherein the incident angle is 10-35 degrees; due to the difference in focal length of the concave mirror 31 in the horizontal and vertical planes, a fluorescence signal (containing the wavelength λ 1 、λ 2 、λ 3 、λ 4 、λ 5 ) Astigmatism is generated in the horizontal and vertical directions after reflection by the concave mirror 31;
fluorescent signal with astigmatism (containing wavelength lambda 1 、λ 2 、λ 3 、λ 4 、λ 5 ) Passes through the cylindrical lens 41 and then is incident on the spectroscope 51. Wherein the convex surface of the cylindrical lens 41 faces the concave mirror, and the convex curvature surface is arranged in the vertical direction, and the cylindrical lens focuses only the fluorescence in the vertical direction, but not the fluorescence in the horizontal direction when the fluorescence signal passes through the cylindrical lens, so that the fluorescence signal (including the wavelength lambda can be corrected 1 、λ 2 、λ 3 、λ 4 、λ 5 ) Astigmatism of (2); meanwhile, the beam splitter 51 is located near the position of twice the focal length of the concave mirror 31, so that the spot sizes in the horizontal and vertical directions of the fluorescent signal on the beam splitter 51 are substantially close and substantially equal to the optical fiber core diameter.
The spectroscope 51 is coated with lambda 1 High transmission, less than lambda 1 A high-reflection wavelength film system; thus wavelength lambda 1 The fluorescence of (2) passes through a spectroscope 51 and has a wavelength lambda 2 、λ 3 、λ 4 、λ 5 Then reflection will occur. Wavelength lambda 1 The fluorescence of (2) passes through the spectroscope 51, the band-pass filter 61 and then is imaged on the photosensitive surface of the detector 81 through the aspherical lens 71. Wherein the photosensitive surface of the detector 81 and the spectroscope 51 are both located near the position of the focal length of the aspherical lens 71, and the aspherical lens 71 can correct the spherical aberration of fluorescence and chromatic aberration, so that the fluorescent light spot on the spectroscope 51 can be imaged on the photosensitive surface of the detector 81 with 1x magnification. The detector 81 may be a photomultiplier tube (PMT) or a small surface area<1 mm) Avalanche Photodiodes (APD).
In short, the wavelength is lambda 1 The fluorescent signal of (2) is outputted from the optical fiber output end face 22, passes through the concave mirror 31, the cylindrical lens 41, the spectroscope 51, the band-pass filter 61, and finally passes through the aspherical lens 71 to be amplified by 1 ×The rate is imaged onto the photosensitive surface of detector 81. And the size of the fluorescent light spot on the photosensitive surface of the detector is basically equal to the core diameter of the optical fiber. And the wavelength is lambda 2 ，λ 3 ，λ 4 ，λ 5 And the fluorescent signal of (c) is reflected by the spectroscope 51.
The fluorescent signal reflected by the spectroscope 51 (including the wavelength lambda 2 、λ 3 、λ 4 、λ 5 ) Transmitted to the concave mirror 32 at a certain incident angle and reflected; wherein, the incident angle is determined by the reflection angle of the fluorescent signal on the spectroscope 51 and is in direct proportion to the numerical aperture NA of the optical fiber 2; wherein the beam splitter 51 is located near the position of the concave mirror 32 at the double focal length; due to the difference in focal length of concave mirror 32 in the horizontal and vertical planes, a fluorescence signal (containing wavelength λ 2 、λ 3 、λ 4 、λ 5 ) Astigmatism is generated after reflection by the concave mirror 32;
fluorescent signal with astigmatism (containing wavelength lambda 2 、λ 3 、λ 4 、λ 5 ) Through the cylindrical lens 42 and then incident on the beam splitter 52. Wherein the convex surface of the cylindrical lens 42 faces the concave mirror 32, and the convex curvature surface is disposed in a vertical direction, and when the fluorescent signal passes through the cylindrical lens 42, the cylindrical lens 42 focuses only the fluorescent signal in the vertical direction, but not the fluorescent signal in the horizontal direction, so that the fluorescent signal (including the wavelength lambda can be corrected 2 、λ 3 、λ 4 、λ 5 ) Astigmatism of (2); meanwhile, the beam splitter 52 is located near the position of the double focal length of the concave mirror 32, so that the spot sizes in the horizontal and vertical directions of the fluorescence on the beam splitter 52 are substantially close to each other, and are substantially equal to the optical fiber core diameter.
The beam splitter 52 is coated with a wavelength lambda 2 High transmission, less than lambda 2 A film system with high reflection of the wavelength; thus wavelength lambda 2 The fluorescence of (2) can pass through the spectroscope 52 and the wavelength is lambda 3 、λ 4 、λ 5 And the fluorescence of (2) will be reflected. Wavelength lambda 2 Which in turn passes through beam splitter 52, bandpass filter 62, and finally is imaged by aspheric lens 72 onto the photosensitive surface of detector 82. Wherein the photosensitive surface of detector 82 and beam splitter 52 are both located on an aspheric lensNear the position of the focal length of 72, the aspheric lens 72 corrects for spherical aberration of fluorescence and chromatic aberration, so that the fluorescent light spot on the beam splitter 52 is imaged onto the photosensitive surface of the detector 72 at 1x magnification. The detector 72 may be a photomultiplier tube (PMT) or a small surface area<1 mm) Avalanche Photodiodes (APD).
In short, the wavelength is lambda 2 The fluorescent signal is output from the optical fiber output end face 22, passes through the concave mirror 31, the cylindrical lens 41, the spectroscope 51, the concave mirror 32, the cylindrical lens 42, the spectroscope 52 and the band-pass filter 62, and finally passes through the aspheric lens 72 to be imaged on the photosensitive surface of the detector 82 at a magnification of 1x, and the fluorescent light spot size on the photosensitive surface of the detector 82 is basically equal to the core diameter of the optical fiber. And lambda is 3 ，λ 4 ，λ 5 And the fluorescent signal is reflected by the beam splitter 52 and continues to be transmitted in the optical path.
Fluorescence is reflected and transmitted back and forth in the concave mirror group and the spectroscope group with the reflecting function, the track is a Z-shaped light path, in the light path, the cylindrical lens corrects astigmatism, the aspherical lens corrects spherical aberration, chromatic aberration and other spherical aberration, and in short:
wavelength lambda 3 The fluorescent signal of (2) is outputted from the optical fiber output end face 22, passes through the concave mirrors 31, 32, the cylindrical lenses 41, 42, the spectroscopes 51, 52, the concave mirror 33, the cylindrical lens 43, the spectroscope 53, the band-pass filter 63 and the aspherical lens 73, and finally is imaged on the photosensitive surface of the detector 83 at a magnification of 1 x. And the size of the fluorescent light spot on the photosensitive surface of the detector 83 is substantially equal to the core diameter of the optical fiber. Spectroscope 53 is coated with lambda 3 High transmission wavelength less than lambda 3 A film system with high reflection of the wavelength; thus lambda 4 ，λ 5 And the fluorescent signal is reflected by the beam splitter 53 and continues to be transmitted in the optical path.
Wavelength lambda 4 The fluorescent signals are output from the optical fiber output end face 22, pass through the concave mirrors 31, 32 and 33, the cylindrical lenses 41, 42 and 43, the spectroscopes 51, 52 and 53, then pass through the concave mirror 34, the cylindrical lens 44 and the spectroscope 54, pass through the band-pass filter 64 and the aspherical lens 74, and finally are imaged on the photosensitive surface of the detector 84 at a magnification of 1 x. And the size of the fluorescent spot on the photosensitive surface of the detector 84 and the fiber coreThe diameters are substantially equal. Beam splitter 54 is coated with lambda 4 High transmission wavelength less than lambda 4 A film system with high reflection of the wavelength; thus lambda 5 And the fluorescent signal is reflected by the beam splitter 54 and continues to be transmitted in the detection system.
Wavelength lambda 5 The fluorescent signals are output from the optical fiber output end face 22, pass through the concave mirrors 31, 32, 33 and 34, the cylindrical lenses 41, 42, 43 and 44, the spectroscopes 51, 52, 53 and 54, then pass through the concave mirror 35, the cylindrical lens 45 and the spectroscope 55, pass through the band-pass filter 65 and the aspherical lens 75, finally are imaged on the photosensitive surface of the detector 85 at 1x magnification, and the fluorescent light spot size on the photosensitive surface of the detector is basically equal to the core diameter of the optical fiber. Spectroscope 55 is coated with lambda 5 High transmission, less than lambda 5 Is a film system with high reflection of the wavelength.
In the above embodiment, the fluorescence signal (including the wavelength lambda) can be realized according to the actual requirement of the customer and the number of wavelengths to be measured 1 、λ 2 、λ 3 、λ 4 And lambda (lambda) 5 ) Is provided.
In the above embodiment, the number of concave mirrors, cylindrical lenses, spectroscopes, band-pass filters, aspheric lenses and detectors can be increased according to the actual demands of customers and the number of wavelengths to be detected, so that the analysis and detection of fluorescent signals with more than 5 wavelengths can be realized, and the number of detected fluorescent wavelengths is not limited by the divergence angle and aberration in transmission.
For the above embodiment, the path track of the fluorescence signal reflected and transmitted back and forth in the concave mirror group and the beam splitter group with reflection function is Z-shaped. The Z-track path is now simplified to fig. 3. FIG. 3 is a schematic view of a concave mirror assembly and a beam splitter assembly of the present invention, wherein the cylindrical lens assembly, the bandpass filter assembly, the lens assembly and the detector assembly of FIG. 2 are omitted. Note that the fluorescent signal output from the optical fiber is first reflected by the concave mirror M (1) and then imaged at the beam splitter D (1), and the concave mirror M (1) of fig. 1 is not shown in fig. 3. Wherein:
a (0): the fluorescent signal light spot can be the aperture of the optical fiber;
a (n): spot diameter at the nth beam splitter;
d (n): an nth spectroscope;
m (n): an nth concave mirror;
in this design, a (0) is a fluorescent signal spot that is equal to the fiber core; after being reflected by a concave mirror M (1), a fluorescent signal light spot A (0) output by an optical fiber is imaged at a spectroscope D (1) to obtain a light spot A (1), then the light spot A (1) is reflected by the concave mirror M (2) and imaged at the spectroscope D (2) to obtain a light spot A (2), and then the light spot A (n-1) at the spectroscope D (n-1) is reflected by the concave mirror M (n) and imaged at the spectroscope D (n) to obtain a light spot A (n). In this design, A (1) is located in front of concave mirror M (2), A (2) is located behind concave mirror M (2), and so on A (n-1) is located in front of concave mirror M (n), A (n) is located behind concave mirror M (n); meanwhile, the dichroic mirror D (1) is located near the position of twice the focal length of the concave mirror M (2), and so on, and the dichroic mirror D (n-1) is located near the position of twice the focal length of the concave mirror M (n).
The angle of incidence of the fluorescence at the concave mirror M (2) is determined by its angle of reflection at D (1) and is proportional to the numerical aperture of the fiber if the magnification from a (0) to a (1) is 1: m, then the magnitude of the angle of incidence at a (1) is equal to the angle of incidence at a (0) multiplied by a factor m. In this design, a (1) to a (2), a (2) to a (3), and so on, the magnifications of a (n) to a (n+1) are all equal to 1, i.e., m is equal to 1. Therefore, the invention can sequentially arrange the fluorescent signals output by the optical fibers according to the wavelength in space and image the fluorescent signals on the spectroscope with the magnification of 1 x. Meanwhile, the spectroscope and the detector are both positioned at the position of the double focal length of the corresponding aspheric lens, so that the whole system can sequentially arrange fluorescent signals output by the optical fiber in space according to wavelength and image the fluorescent signals on the photosensitive surface of the detector at 1X magnification, and the size of fluorescent light spots on the photosensitive surface of the detector is basically equal to the core diameter of the optical fiber.
In summary, after the fluorescent signals with multiple wavelengths are incident to the concave mirror through the optical fiber, the fluorescent signals are sequentially separated in space through the cylindrical lens group and the spectroscope group; in the transmission process, a cylindrical mirror corrects the astigmatism of a fluorescent signal, an aspherical mirror corrects the aberration such as spherical aberration and chromatic aberration, and the fluorescent signal is orderly arranged in space according to the wavelength and is incident on a detector with corresponding wavelength at 1 multiplied by the amplification rate; the number of fluorescence wavelengths detected by system analysis can be increased or decreased according to the needs of clients, so that the system has better client adaptability; meanwhile, the optical device has low requirement on adjustment precision, and has better cost advantage and production process.
The foregoing is a further detailed description of the invention in connection with specific embodiments, and it is not intended that the invention be limited to such description. It will be apparent to those skilled in the art that several simple deductions or substitutions may be made without departing from the spirit of the invention, and these should be considered to be within the scope of the invention.
Claims (10)
1. A fluorescence detection system for a cellular analyzer, characterized by: the fluorescent detection system comprises an objective lens, an optical fiber, a concave lens group, a cylindrical lens group, a spectroscope group, a band-pass filter group and a detector group which form a detection area, and the fluorescent detection system formed by the components collects and detects fluorescence emitted by dyed sample particles passing through the detection area after laser irradiation;
wherein:
the objective lens is used for collecting and collimating fluorescence generated after the sample particles are excited by the laser;
the optical fiber comprises an incident end face and an emergent end face, and is used for transmitting fluorescent light collimated by the objective lens;
the concave mirror group comprises a plurality of concave mirrors which are arranged in an array and are used for reflecting fluorescence, the concave surfaces of the concave mirrors are plated with total reflection films, and the concave surfaces of the concave mirrors face to the optical fibers;
the cylindrical lens group comprises a plurality of planar convex cylindrical lenses which are arranged in an array and used for correcting astigmatism, and the planar convex cylindrical lenses are used for correcting astigmatism, and both surfaces of each cylindrical lens are plated with an antireflection film system;
the spectroscope group comprises a plurality of spectroscopes which are arranged in an array, each spectroscope is respectively coated with a film according to the fluorescent wavelength to be detected, so that fluorescent signals larger than a certain wavelength pass through, and fluorescent signals smaller than the wavelength are reflected;
the band-pass filter set comprises a plurality of band-pass filters which are arranged in an array, each filter is coated with a film according to the wavelength of fluorescence to be detected, so that fluorescence with a specific wavelength passes through, and fluorescence with other wavelengths is totally reflected;
the lens group comprises a plurality of aspherical lenses which are arranged in an array, two surfaces of each aspherical lens are plated with an antireflection film system, and the aspherical lenses image fluorescent light spots on the spectroscope on a photosensitive surface of the detector;
the detector group comprises a plurality of detectors which are arranged in an array, the detectors are arranged corresponding to the fluorescent wavelengths divided by the band-pass filter, and the detectors are used for receiving fluorescent signals and converting the fluorescent signals into electric signals.
2. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the spectroscope is arranged near the position of the double focal length of the concave mirror corresponding to the spectroscope.
3. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the photosensitive surface of the detector is positioned near the position of the double focal length of the corresponding aspheric lens.
4. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the spectroscope is positioned near the position of the double focal length of the corresponding aspheric lens.
5. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the incident end face of the optical fiber is positioned near the focal point of the objective lens; the emergent end face of the optical fiber is positioned near the position of the double focal length of the concave mirror.
6. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the objective lens has a high Numerical Aperture (NA) of 0.5-1.4.
7. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the optical fiber has a relatively low numerical aperture and NA of 0.05-0.15.
8. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the aspherical lenses arranged in an array have higher numerical aperture and NA of 0.5-0.9.
9. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the convex curvature surfaces of the cylindrical lenses arranged in an array are opposite to the detector.
10. A fluorescence detection system for a cellular analyzer as defined in claim 1, wherein: the photosensitive surfaces of the detectors arranged in an array face the lens; the detector is a photomultiplier
(PMT) or Avalanche Photodiode (APD).
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| CN110361074B (en) * | 2018-03-26 | 2020-09-08 | 华中科技大学 | Photoelectric liquid level detection device |
| WO2020075293A1 (en) * | 2018-10-12 | 2020-04-16 | 株式会社日立ハイテクノロジーズ | Dichroic mirror array and light detecting device |
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