CN116500004A - A small fluorescence analysis optical system and method for microfluidic chip detection - Google Patents

Abstract

The invention relates to a small-sized fluorescence analysis optical system and a method for detecting a microfluidic chip. The system comprises a laser, a first small aperture diaphragm, a first optical filter, a dichroic mirror, an optical trap, a microfluidic chip, a mask, a collimating lens, a condensing lens, a second optical filter, a second small aperture diaphragm and a photoelectric detector. The invention improves the light path structure layout and reduces the whole volume of the optical system by the light path miniaturized structure design while ensuring the detection stability and the sensitivity; the dichroic mirror and the optical trap are arranged on the optical path, so that secondary filtering is carried out on the excitation light, stray light is effectively reduced, and interference of background noise in the detection process of the oblique optical system is reduced; the mask is arranged on the surface of the microfluidic chip, so that autofluorescence phenomenon generated after the microfluidic chip is irradiated by excitation light can be effectively inhibited, and scattering of the excitation light on the surface of the detection area of the microfluidic chip is reduced; and the micro-fluidic chip is subjected to scanning detection, so that quantitative analysis of the low-concentration sample to be detected is realized.

Description

A small fluorescence analysis optical system and method for microfluidic chip detection

technical field

The invention relates to the technical field of laser-induced fluorescence detection of biological immunofluorescence analysis and the technical field of microfluidic chip in vitro diagnosis, in particular to a small fluorescence analysis optical system and method for microfluidic chip detection.

Background technique

Early diagnosis, treatment and curative effect monitoring of diseases are of great significance to the promotion of human health. Effective treatment of many critical illnesses depends on the timeliness and accuracy of diagnosis. However, traditional detection methods such as tissue biopsy still have limitations such as difficulty in sampling, failure to achieve early detection, pain for patients, and low sampling frequency. With the development of biochemistry, immunology, molecular biology and other fields, as well as the rise of molecular diagnosis and precision medicine, in vitro diagnostic technology has made remarkable achievements in the application of molecular diagnosis and point-of-care detection.

Immunofluorescence analysis has a wide range of detection types, and has the advantages of strong specificity, high sensitivity, and good practicability. It is one of the commonly used detection techniques in the field of in vitro diagnostics. Immunofluorescence analysis is often used to measure biologically active compounds with very low levels, such as proteins (enzymes, receptors, antibodies), hormones (steroids, thyroid hormones, phthalotropic hormones), cells and microorganisms, etc. Qualitative and quantitative analysis of the biological activity of the target. The traditional immunofluorescence analysis system has the advantages of high precision and high-throughput detection, but there are problems such as large system volume, high price, complicated operation, large reagent consumption, and long detection time, making it difficult to achieve rapid and real-time detection. At the same time, this type of system is mainly used in hospitals, university laboratories, and scientific research institutions. It has high requirements for experimental environment, sample pretreatment, and professional operation capabilities, and cannot be popularized for daily use.

With the rise of microfluidic chip technology, the biochemical experiment platform is built on the microfluidic chip, which has the advantages of small system size, high real-time detection, low comprehensive cost, and does not rely on professional equipment, etc., and can be widely used in clinical monitoring , inspection and quarantine, family health care and other fields, its rapid development conforms to the efficient and fast-paced working methods of the current society, meets people's needs in terms of time, and enables patients to receive effective diagnosis and treatment as soon as possible.

The light path structure of the small immunofluorescence analysis system used for microfluidic chip detection can be mainly divided into four types, including: confocal type, oblique type, orthogonal type and parallel type. Compared with other types of optical systems, the oblique optical system has the characteristics of simple optical path structure, small size, and low manufacturing cost, and is more suitable for the research of small portable diagnostic instruments. The optical path principle of the traditional oblique optical system is shown in Figure 1. The excitation light emitted by the light source is first filtered by a short-pass filter, then condensed by a collimator lens, and irradiates the detection area of the microfluidic chip at an appropriate incident angle. , the target to be measured marked with fluorescein on the detection area is excited by the laser to generate fluorescence, which is collected by the condenser lens, and the excitation light and stray light are filtered out by the long-wave pass filter, and finally collected by the photodetector. However, there are also problems with the oblique optical system: the optical system is easily affected by excitation light and stray light during the detection process, resulting in low detection sensitivity. Therefore, it is particularly important to effectively reduce background noise interference to improve the detection sensitivity of the optical system.

Contents of the invention

The object of the present invention is to provide a small fluorescence analysis optical system and method for microfluidic chip detection, the optical system and method can solve the deficiencies in the prior art. At the same time, a mask structure is added to reduce the background noise in the detection area, and a microfluidic chip composed of polymethyl methacrylate (PMMA) and paper-based detection unit is scanned to realize the detection of low-concentration biological samples detection.

To achieve the above object, the present invention adopts the following technical solutions:

In the first aspect of the present invention, a small fluorescence analysis optical system for microfluidic chip detection is disclosed, the system includes:

The laser emission processing optical path is used to emit laser light, shape and filter the emitted laser light, and focus the laser light in the target band to the detection area of the microfluidic chip;

Microfluidic chip, after the laser is irradiated to the detection area of the microfluidic chip, the sample to be tested captured by the paper-based detection unit in the detection area is excited by the laser of the target band to release fluorescence;

The fluorescence collection and detection optical path is used to collect the fluorescent signal released on the detection area of the microfluidic chip, and obtain the fluorescence distribution curve of the sample to be tested after the immunofluorescence reaction by scanning the detection area.

Further, the optical path for laser emission processing includes:

a laser, a first pinhole diaphragm, a first filter, a dichroic mirror and a light trap;

The laser is placed vertically and horizontally, forming an angle of 90° with the microfluidic chip;

The first aperture diaphragm is arranged below the laser and placed parallel to the horizontal plane;

The first optical filter is arranged below the first aperture diaphragm and placed parallel to the horizontal plane;

The dichroic mirror is arranged below the first filter and forms an included angle of 60° with the projection of the horizontal plane;

The light trap is disposed below the dichroic mirror and parallel to the horizontal plane.

Further, the fluorescence collection and detection light path includes:

A microfluidic chip, a detection area, a mask, a collimating lens, a condenser lens, a second filter, a second aperture diaphragm and a photodetector;

The microfluidic chip is arranged below the mask and placed parallel to the horizontal plane;

The mask is arranged below the dichroic mirror and placed parallel to the horizontal plane;

The mask is arranged above the microfluidic chip and placed parallel to the horizontal plane;

The collimating lens is arranged above the microfluidic chip and placed parallel to the horizontal plane;

The condenser lens is arranged above the collimator lens and placed parallel to the horizontal plane;

The second filter is arranged above the condenser lens and placed parallel to the horizontal plane;

The second aperture diaphragm is arranged above the second filter and placed parallel to the horizontal plane;

The photodetector is arranged above the second aperture diaphragm and placed vertically to the horizontal plane.

Further, the dichroic mirror is used to transmit the stray light that is not completely filtered out in the laser, the stray light is absorbed by the mask, and the dichroic mirror is used to reflect the excitation light of the target wavelength band, The reflected excitation light forms an included angle of 30° with its projection on the horizontal plane.

Further, the projection of the dichroic mirror and the horizontal plane forms an included angle of 60°.

Further, the microfluidic chip includes a microfluidic chip body and a microfluidic channel provided on the microfluidic chip body. The main body of the microfluidic chip on both sides of the microfluidic channel is provided with grooves for serving as boundaries of the microfluidic channel. The micro-channel is provided with a detection area, and the detection area is provided with several grooves, and paper-based detection units are placed in the grooves; specifically, two square grooves are arranged in the middle of the detection area, A paper-based detection unit is respectively placed in the two square grooves, and the two paper-based detection units serve as the detection line (T line) and the quality control line (C line) of the detection area respectively. The capture antibody is immobilized on the paper-based detection unit for capturing the sample to be tested in the sample reagent.

Further, the main body of the microfluidic chip is made of polymethyl methacrylate material;

The paper-based detection unit is made of nitrocellulose membrane.

Further, the mask covers the upper surface of the microfluidic chip;

The mask is provided with a scanning window above the detection area.

In the second aspect of the present invention, an analysis method of the above-mentioned fluorescence analysis optical system is disclosed, the method includes:

The laser emission processing optical path is used to emit laser light, and the emitted laser light is shaped and filtered to focus the laser light in the target band to the detection area of the microfluidic chip;

The paper-based detection unit on the detection area of the microfluidic chip is used to capture and fix the immunofluorescence-labeled sample to be tested, and release fluorescence after excitation of the laser in the target band;

The fluorescent signal released on the detection area of the microfluidic chip is collected by the fluorescence collection and detection optical path, and the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested is obtained by scanning the detection area.

Further, the laser emission processing optical path is used to emit laser light, and the emitted laser light is shaped and filtered to focus the laser light in the target band onto the detection area of the microfluidic chip, including:

The laser light emitted by the laser is firstly shaped into a circular spot through the first small hole aperture. After the laser passes through the first optical filter to filter the stray light, the dichroic mirror passes through the stray light that has not been completely filtered out in the laser. The trap collects, and reflects the excitation light of the target wavelength band to the detection area of the microfluidic chip, forming a spot on the detection area.

Further, the paper-based detection unit on the detection area of the microfluidic chip is used to capture and fix the immunofluorescence-labeled sample to be tested, and release fluorescence after the excitation of the laser in the target band, including:

Place the microfluidic chip that has completed the immunofluorescent labeling of the sample to be tested at the detection position, start the laser, and the sample to be tested that is labeled with fluorescein in the detection area of the microfluidic chip is excited by the excitation light and releases fluorescence.

Further, the fluorescence collection and detection optical path is used to collect the fluorescent signal released on the detection area of the microfluidic chip, and by scanning the detection area, the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested is obtained, including:

Fluorescence is collimated and focused by a condenser lens group composed of a collimator lens and a condenser lens, and stray light is blocked by a second filter to filter the background light and a second aperture diaphragm;

By irradiating the photosensitive surface of the photodetector with fluorescence, the photodetector converts the collected fluorescent signal into an electrical signal;

The stage is driven by a stepping motor to drive the microfluidic chip to perform step-by-step operation in one direction. The optical system remains fixed, and the photodetector collects one data at each step of operation, and gradually scans from left to right in the detection area to collect multiple data. The data are used to obtain the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested on the microfluidic chip.

Compared with prior art, the advantages of the present invention are:

(1) Through the miniaturized structure design of the optical path, the present invention enables the oblique laser-induced fluorescence detection optical path to ensure the stability and sensitivity of the detection, improve the structural layout of the optical path, reduce the overall volume of the optical system, and make the optical system more suitable for portable rapid diagnosis System development.

(2) The present invention can effectively reduce the stray light in the excitation light by arranging dichroic mirrors and optical traps on the optical path to filter the excitation light twice, and reduce the interference of background noise in the detection of oblique optical systems. Greater signal-to-noise ratio is achieved in system fluorescence detection.

(3) The present invention can effectively suppress the autofluorescence phenomenon produced after the microfluidic chip is irradiated by excitation light by setting a mask on the surface of the microfluidic chip, and reduce the scattering of the excitation light on the surface of the detection area of the microfluidic chip , to improve the sensitivity of the oblique optical system to detect fluorescent signals.

(4) The present invention uses a combination of a microfluidic chip and an optical system for fluorescence detection and analysis, wherein the microfluidic chip is made of polymethyl methacrylate (PMMA) material as the main body of the microfluidic chip, and is made of nitrocellulose membrane The paper-based detection unit is used as the test line (T line) and quality control line (C line) of the detection area in the microfluidic channel on the microfluidic chip. While the microfluidic chip realizes the stable flow delay of reagent samples, the paper-based The detection unit can improve the detection efficiency of the fluorescent signal, and is beneficial to the detection of low-concentration samples to be tested.

(5) The present invention realizes the collection of fluorescence data of the sample to be tested on the microfluidic chip through the oblique optical system, uses the optical system to scan the detection area of the microfluidic chip, and uses the photodetector to convert the photoelectric signal to obtain the fluorescence distribution curve , which can clearly reflect the most original situation of the fluorescence of the sample to be tested on the microfluidic chip, and provide strong basic data for studying the fluorescence excitation of the sample to be tested at low concentration.

Description of drawings

Fig. 1 is a schematic diagram of the principle of an existing oblique beam optical system;

Fig. 2 is the schematic diagram of the principle of the optical system in the present invention;

Fig. 3 is a schematic structural view of the optical system in the present invention;

Fig. 4 is a scanning diagram of fluorescence signals detected by the optical system on the microfluidic chip in the present invention.

in:

a1, light source; a2, short-wave pass filter; a3, collimating lens in the existing oblique optical system; a4, microfluidic chip in the existing oblique optical system; a5, in the existing oblique optical system a6, long-wave pass filter; a7, the photodetector in the existing oblique optical system; 1, laser; 2, the first aperture stop, 3, the first filter; 4, Dichroic mirror; 5, optical trap; 6, microfluidic chip; 7, detection area; 8, mask; 9, collimating lens; 10, condenser lens; 11, second filter; 12, second 2. Small hole diaphragm; 13. Photoelectric detector.

Detailed ways

The present invention will be further described below in conjunction with accompanying drawing:

In order to solve the problem of the immunofluorescence analysis system used for microfluidic chip detection, the traditional optical system is large in size, which is not conducive to integration in the portable immunofluorescence diagnostic device. For problems such as low detection sensitivity of samples, the present invention provides a small-scale fluorescence analysis optical system and method for microfluidic chip detection. A mask structure is added to reduce background noise in the detection area, and a microfluidic chip composed of polymethyl methacrylate (PMMA) and paper-based detection units is scanned to realize the detection of low-concentration biological samples to be tested.

As shown in Figures 2 and 3, a small fluorescence analysis optical system for microfluidic chip detection, the optical system includes a laser emission processing optical path, a microfluidic chip 6, and a fluorescence collection and detection optical path. The laser emission processing optical path is used to emit laser light and shape and filter the emitted laser light, and focus the laser light in the target band onto the detection area 7 of the microfluidic chip 6; After the detection area of the chip, the sample to be tested captured by the paper-based detection unit in the detection area 7 releases fluorescence after being excited by the laser in the target band; the fluorescence collection and detection optical path is used to collect the fluorescence released on the detection area 7 of the microfluidic chip 6 signal, by scanning the detection area 7 to obtain the fluorescence distribution curve of the sample to be tested after the immunofluorescence reaction.

Specifically, the optical path for laser emission processing includes a laser 1 , a first aperture stop 2 , a first filter 3 , a dichroic mirror 4 and an optical trap 5 arranged in sequence. The optical path for fluorescence collection and detection includes a mask 8 , a collimator lens 9 , a condenser lens 10 , a second filter 11 , a second aperture stop 12 and a photodetector 13 arranged in sequence. The laser 1 is placed vertically to the horizontal plane, forming an angle of 90° with the microfluidic chip 6 . The first aperture diaphragm 2 is arranged below the laser 1 and placed parallel to the horizontal plane. The first filter 3 is arranged below the first aperture stop 2 and placed parallel to the horizontal plane. The dichroic mirror 4 is disposed under the first filter 3 and forms an included angle of 60° with the projection of the horizontal plane. The light trap 5 is disposed below the dichroic mirror 4 and parallel to the horizontal plane. The microfluidic chip 6 is arranged under the mask 8 and placed parallel to the horizontal plane. The detection area 7 is arranged on the microfluidic channel of the microfluidic chip 6, and the detection area 7 is provided with two square grooves, and the paper-based detection unit is placed in the square grooves; the mask 8 is arranged on the two square grooves. Below the dichroic mirror 4, it is placed parallel to the horizontal plane. The mask 8 is arranged above the microfluidic chip 6 and placed parallel to the horizontal plane. The collimating lens 9 is arranged above the microfluidic chip 6 and placed parallel to the horizontal plane. The condenser lens 10 is arranged above the collimator lens 9 and placed parallel to the horizontal plane. The second filter 11 is disposed above the condenser lens 10 and placed parallel to the horizontal plane. The second aperture diaphragm 12 is disposed above the second filter 11 and placed parallel to the horizontal plane. The photodetector 13 is arranged above the second aperture diaphragm 12 and placed perpendicular to the horizontal plane.

Further, on both sides of the microfluidic channel on the main body of the microfluidic chip 6, a groove of 1.0*1.0mm (depth*width) is respectively provided as the boundary of the microfluidic channel, and the height of the microfluidic channel is 0.15mm. The overall size is 42*16*4.0mm (length*width*height).

Further, the size of the detection area 7 in the microfluidic channel on the microfluidic chip 6 is 10*2.0mm

(length*width). Two square grooves with a size of 2.0*2.0*0.3mm (length*width*depth) are set in the middle of the detection area 7 as grooves for placing paper-based detection units, and the interval between the two square grooves is 2mm. Two square slots are respectively used to place a paper-based detection unit with a size of 2.0*2.0*0.3mm (length*width*height), and the two paper-based detection units are respectively used as the test line (T line) of the detection area 7 And quality control line (C line). The paper-based detection unit on the detection area 7 is used to capture the sample to be tested in the sample reagent by immobilizing the capture antibody.

Further, the main body of the microfluidic chip 6 is made of polymethyl methacrylate (PMMA) and other low-cost, easy-to-machine materials, and the paper-based detection unit in the detection area 7 uses a nitrocellulose membrane, etc. A type of paper material.

Further, the laser 1 is a red light semiconductor laser, and the center wavelength of the emitted laser light is 660nm. A focusing lens is set to adjust the size of the laser spot, so that the laser light emitted by the laser 1 is irradiated on the detection area 7 of the microfluidic chip 6 The diameter of the spot on is 2.0mm.

Further, the projection of the dichroic mirror 4 and the horizontal plane is at an angle of 60°, the transmission wavelength range is 665-800nm, and the reflection wavelength range is 610-660nm, which is used to transmit the stray light that is not completely filtered out in the laser, The stray light is absorbed by the mask 8, and the dichroic mirror 4 is used to reflect the excitation light of the target wavelength band, and the projection of the reflected light on the horizontal plane forms an included angle of 30°. The transmission band of the dichroic mirror 4 is 665-800nm, and the reflection band is 610-660nm.

Further, the mask 8 covers the upper surface of the microfluidic chip 6 with a size of 42*16*0.5mm (length*width*height), and the mask 8 is in the detection area of the microfluidic chip 6 A scanning window of 18*2.5mm is set above 7, so that the test line (T line) and the quality control line (C line) of the detection area 7 are located in the middle of the scanning window of the mask 8. The mask 8 uses a light-absorbing material such as flocked light-absorbing black cloth, which is used to suppress the autofluorescence phenomenon generated after the microfluidic chip 6 is irradiated by excitation light, and reduce the excitation on the surface of the detection area 7 of the microfluidic chip 6. light scattering.

Further, the aperture of the first aperture stop 2 is 1.0 mm, which is used to shape the laser light emitted by the laser 1, so that the laser light is irradiated on the detection area 7 of the microfluidic chip 6 to form a circle. shaped spot.

Further, the light trap 5 uses light-absorbing materials such as flocked light-absorbing black cloth to absorb stray light that is not completely filtered out in the laser.

In this example, using the fluorescein Alexa 680 Marking and staining of the samples to be tested, fluorescein Alexa/> The absorption peak wavelength of 680 is 679nm, the emission peak wavelength is 702nm, fluorescein Alexa/> The fluorescence signal excited by 680 is in the near-infrared light band, which can effectively avoid the noise interference caused by the external visible light band. It is a commonly used quantitative analysis method for detecting protein targets; The wavelength is 665nm, the bandwidth is 20nm, the center wavelength of the second optical filter 11 is 715nm, the bandwidth is 20nm; optional, according to the need to detect different types of fluorescent wavelengths, the optical system in this embodiment can be replaced with a matching optical characteristic Lasers, filters.

Most traditional small optical systems use only one condenser to collect fluorescence. The nature of the condenser determines that the light collection characteristics of a single condenser are not high. In this embodiment, a combination of double condensers is used to improve the fluorescence focusing efficiency. Wherein, the collimating lens 9 is a biconvex lens with a diameter of 12.7 mm, the focal length is 12.5 mm, and the back focus is 10.51 mm; the condenser lens 10 is a single convex lens with a diameter of 12.7 mm, the focal length is 15.0 mm, and the back focus is 11.63 mm. The aperture diameter of the second aperture diaphragm 12 can be selected from 1.0-2.0 mm, which is optimized according to the photosensitive surface of the photodetector 13 . The photodetector 13 is a photodiode (PD) or an avalanche photodiode (APD), which is used to convert the received fluorescent signals of different amplitudes into electrical signals, so as to count the samples to be tested in the detection area of the microfluidic chip 6. Fluorescent signal curves corresponding to the detection line (T line) and quality control line (C line) in 7.

The analysis method of the above-mentioned small-scale fluorescence analysis optical system used for the detection of the microfluidic chip 6 is:

Place the microfluidic chip 6 that has completed the immunofluorescent labeling of the sample to be tested on the detection position of the optical system, start the laser 1, and the laser light emitted by the laser 1 first passes through the first small hole diaphragm 2 and is shaped into a circular spot, and the laser passes through the second aperture. After a filter 3 filters the stray light, the dichroic mirror 4 passes through the stray light that has not been completely filtered out in the laser, and collects it by the optical trap 5, and reflects the laser light of the target band to the detection area 7 of the microfluidic chip 6 , forming a light spot with a diameter of 2.0 mm on the detection area 7 .

The sample to be tested labeled with fluorescein in the detection area 7 of the microfluidic chip 6 is excited by the excitation light and releases fluorescence. The fluorescence is collimated and focused by the condenser lens group composed of the collimator lens 9 and the condenser lens 10. After the first The second optical filter 11 filters the background light and the second aperture diaphragm 12 blocks stray light, and irradiates the photosensitive surface of the photodetector 13 through the fluorescence, and the photodetector 13 converts the collected fluorescence signal into an electrical signal.

During the detection process, the stepping motor drives the stage to drive the microfluidic chip 6 to perform stepping operation in one direction, each stepping displacement is 0.04 mm, the optical system remains fixed, and the photodetector 13 collects one data at each step of operation. On the detection area 7 with a length of 10 mm, it scans step by step from left to right, and collects 250 data in total. The fluorescent signal scanning diagram of the optical system of the present invention is shown in Figure 4, which is used to obtain the immune response of the sample to be tested on the microfluidic chip. Fluorescence distribution curve after fluorescence reaction.

The above-mentioned embodiments are only descriptions of preferred implementations of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, those skilled in the art may make various modifications to the technical solutions of the present invention. and improvements, all should fall within the scope of protection determined by the claims of the present invention.

Claims (10)

1. A small fluorescence analysis optical system for microfluidic chip detection, characterized in that the system comprises: The laser emission processing optical path is used to emit laser light, shape and filter the emitted laser light, and focus the laser light in the target band to the detection area of the microfluidic chip; Microfluidic chip, after the laser is irradiated to the detection area of the microfluidic chip, the sample to be tested captured by the paper-based detection unit in the detection area is excited by the laser of the target band to release fluorescence; The fluorescence collection and detection optical path is used to collect the fluorescent signal released on the detection area of the microfluidic chip, and obtain the fluorescence distribution curve of the sample to be tested after the immunofluorescence reaction by scanning the detection area. 2. The system of claim 1, wherein: The optical path for laser emission processing includes: a laser, a first pinhole diaphragm, a first filter, a dichroic mirror and a light trap; The laser is placed vertically and horizontally, forming an angle of 90° with the microfluidic chip; The first aperture diaphragm is arranged below the laser and placed parallel to the horizontal plane; The first optical filter is arranged below the first aperture diaphragm and placed parallel to the horizontal plane; The dichroic mirror is arranged below the first filter and forms an included angle of 60° with the projection of the horizontal plane; The light trap is disposed below the dichroic mirror and parallel to the horizontal plane. 3. The system of claim 1, wherein: The fluorescence collection and detection light path includes: Mask, collimating lens, condenser lens, second optical filter, second aperture diaphragm and photodetector; The microfluidic chip is arranged below the mask and placed parallel to the horizontal plane; The mask is arranged below the dichroic mirror and placed parallel to the horizontal plane; The mask is arranged above the microfluidic chip and placed parallel to the horizontal plane; The collimating lens is arranged above the microfluidic chip and placed parallel to the horizontal plane; The condenser lens is arranged above the collimator lens and placed parallel to the horizontal plane; The second filter is arranged above the condenser lens and placed parallel to the horizontal plane; The second aperture diaphragm is arranged above the second filter and placed parallel to the horizontal plane; The photodetector is arranged above the second aperture diaphragm and placed vertically to the horizontal plane. 4. The system of claim 3, wherein: The dichroic mirror is used to pass through the stray light that is not completely filtered out in the laser, and the stray light is absorbed by the mask, and the dichroic mirror is used to reflect the excitation light of the target band, and the reflected The excitation light forms an angle of 30° with its projection on the horizontal plane. 5. The system of claim 4, wherein: The projection of the dichroic mirror and the horizontal plane forms an included angle of 60°. 6. The system of claim 1, wherein: The microfluidic chip includes a microfluidic chip body and a microfluidic channel arranged on the microfluidic chip body; A detection area is set on the micro-channel, and several grooves are set in the detection area, and paper-based detection units are placed in the grooves; The main body of the microfluidic chip on both sides of the microfluidic channel is provided with grooves for serving as boundaries of the microfluidic channel. 7. The system of claim 6, wherein: The main body of the microfluidic chip is made of polymethyl methacrylate material; The paper-based detection unit is made of nitrocellulose membrane. 8. The system of claim 3, wherein: The mask covers the upper surface of the microfluidic chip; The mask is provided with a scanning window above the detection area. 9. An analysis method of the fluorescence analysis optical system according to any one of claims 1 to 8, characterized in that the method comprises: The laser emission processing optical path is used to emit laser light, and the emitted laser light is shaped and filtered to focus the laser light in the target band to the detection area of the microfluidic chip; The paper-based detection unit on the detection area of the microfluidic chip is used to capture and fix the immunofluorescence-labeled sample to be tested, and release fluorescence after excitation of the laser in the target band; The fluorescent signal released on the detection area of the microfluidic chip is collected by the fluorescence collection and detection optical path, and the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested is obtained by scanning the detection area. 10. The method of claim 9, wherein, The laser emission processing optical path is used to emit laser light, and the emitted laser light is shaped and filtered to focus the laser light in the target band onto the detection area of the microfluidic chip, including: The laser light emitted by the laser is firstly shaped into a circular spot through the first small hole diaphragm. After the laser passes through the first filter to filter the stray light, the dichroic mirror passes through the stray light that has not been completely filtered out in the laser. The trap is collected, and the laser light of the target band is reflected to the detection area of the microfluidic chip, and a spot is formed on the detection area; The paper-based detection unit on the detection area of the microfluidic chip is used to capture and fix the immunofluorescence-labeled sample to be tested, and release fluorescence after the excitation of the laser in the target band, including: Place the microfluidic chip that has completed the immunofluorescent labeling of the sample to be tested at the detection position, start the laser, and the sample to be tested that is labeled with fluorescein in the detection area of the microfluidic chip releases fluorescence after being excited by the laser of the target band; The fluorescence collection and detection optical path is used to collect the fluorescent signal released on the detection area of the microfluidic chip, and the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested is obtained by scanning the detection area, including: Fluorescence is collimated and focused by a condenser lens group composed of a collimator lens and a condenser lens, and stray light is blocked by a second filter to filter the background light and a second aperture diaphragm; By irradiating the photosensitive surface of the photodetector with fluorescence, the photodetector converts the collected fluorescent signal into an electrical signal; The stage is driven by a stepping motor to drive the microfluidic chip to perform step-by-step operation in one direction. The optical system remains fixed, and the photodetector collects one data at each step of operation, and gradually scans from left to right in the detection area to collect multiple data. The data are used to obtain the fluorescence distribution curve after the immunofluorescence reaction of the sample to be tested on the microfluidic chip.