CN108387505B - Multifunctional optical tweezers system and method based on micro-fluidic chip - Google Patents
Multifunctional optical tweezers system and method based on micro-fluidic chip Download PDFInfo
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Abstract
The invention discloses a multifunctional optical tweezers system and a method based on a micro-fluidic chip, wherein the system comprises: the micro-fluidic chip particle sampling system, the counting system, the signal detection system and the optical tweezers sorting system; the micro-fluidic chip particle sample introduction system is used for leading particles to pass through the micro-fluidic chip one by one along a certain path through hydrodynamic focusing; when the particles pass through the detection counting area, the signal detection system realizes simultaneous representation of multiple parameters of a luminescent signal and a forward scattering light signal of the particles by a weak light signal detection technology, realizes detection of the particles, and counts the number of the particles by the counting system; when the particles pass through the capture sorting area, the specific types of particles are deflected by the optical tweezers sorting system according to the detection result of the particles, so that the sorting function of the particles is realized. The device has compact structure, is easy to realize miniaturization, and has the advantages of high sensitivity, high analysis speed, high flux, good selectivity, small sample consumption, strong anti-interference capability and the like.
Description
Technical Field
The invention relates to the cross technical field of microfluidic chip laboratories, optical tweezers micromanipulation technology, nanophotonics, biomedical detection and the like, in particular to a multifunctional optical tweezers system and a method based on a microfluidic chip.
Background
Commonly used cell sorting methods include cell screening, centrifugal sorting, laser-induced fluorescence sorting and magnetic sorting. The commercialized flow cytometry generally adopts laser-induced fluorescence detection and combines a dielectrophoresis force sorting method, and the flow cytometry based on a microfluidic chip can popularize the traditional cell sorting method to the microfluidic technical level for realization, and utilizes the action of external force to realize sorting, such as dielectrophoresis sorting, magnetic adsorption, optical sorting and acoustic sorting, so that the characteristics of small size, high efficiency, high integration level, high analysis speed, low price and the like of the microfluidic chip are fully embodied.
The concept of optical tweezers was first proposed by american scientist Ashkin in 1970, where a highly focused gaussian laser beam produced a strong enough optical trap gradient force at the focal point to trap and manipulate micron and even nanometer sized particles. The optical tweezers technology adopts a non-contact remote control working mode, does not need to process a micro-control part, and does not generate mechanical damage to controlled cells, so that the optical tweezers becomes the most common means for optically controlling single cells or other particles in a micro-fluidic chip.
Upconversion luminescent nanomaterials absorb two or more low energy photons and radiate a high energy photon, typically by converting near infrared light into visible light. It has many advantages as a biomarker probe, for example: low toxicity, high chemical stability, excellent photostability, narrow-band emission, long luminescence lifetime, high signal-to-noise ratio, and the like; in addition, near-infrared lasers offer many advantages as their excitation light sources, such as: the deep light penetration depth has almost no damage to biological tissues, no background fluorescence and the like.
The microsphere-based suspension chip technology is that an antibody or other biomolecules are immobilized on a microsphere carrier to prepare capture microspheres, then the capture microspheres in a suspension state specifically recognize different substances to be detected in a detection system, and finally the capture microspheres are hybridized or immunoreactive with a report antibody or biomolecules to form a sandwich structure. The concentration of the target is quantified by detecting the signal of the reporter molecule on the individual microspheres, such as fluorescence, radioactivity, chemiluminescence, etc. The simultaneous detection of multiple analytes can be realized through the size coding, the fluorescence coding and the like of the microspheres, and the technology is very suitable for the detection of trace samples.
Based on a microfluidic chip and an up-conversion nano-labeling technology, the constructed optical tweezers sorting system has very good application potential in the fields of biological analysis and the like, can carry out high-sensitivity detection on various biomolecules such as nucleic acid, protein and the like or virus particles and other objects to be detected in a complex sample (such as whole serum and whole plasma), and can carry out detection, sorting and typing on Circulating Tumor Cells (CTCs) in a blood sample of a tumor patient, thereby providing a new analysis technology for early diagnosis, curative effect evaluation and cancer metastasis mechanism research of cancer.
Disclosure of Invention
The invention aims to solve the technical problem of providing a multifunctional optical tweezers system and a method based on a micro-fluidic chip aiming at the defects in the prior art.
The technical scheme adopted by the invention for solving the technical problems is as follows:
the invention provides a multifunctional optical tweezers system based on a microfluidic chip, which comprises: the micro-fluidic chip particle sampling system, the counting system, the signal detection system and the optical tweezers sorting system; wherein:
the micro-fluidic chip particle sample introduction system is used for leading particles to pass through the micro-fluidic chip one by one along a certain path through hydrodynamic focusing, and a detection counting area and a capture sorting area are preset on the micro-fluidic chip;
when the particles pass through the detection counting area, the signal detection system realizes simultaneous representation of multiple parameters of a luminescent signal and a forward scattering light signal of the particles by a weak light signal detection technology, realizes detection of the particles, and counts the number of the particles by the counting system;
when the particles pass through the capture sorting area, the specific types of particles are deflected by the optical tweezers sorting system according to the detection result of the particles, so that the sorting function of the particles is realized.
Furthermore, the micro-fluidic chip is provided with 3 paths of inflow ends and 3 paths of outflow ends, and the detection counting area and the capture sorting area are arranged between the inflow ends and the outflow ends; wherein:
in the 3-way inflow end, sheath flow is arranged at two sides, and a sample flow of particles is arranged in the middle; the sample flow and the sheath flow simultaneously flow into the detection counting area and the capture sorting area, the sheath flows at two sides are used for ensuring that the sample flow forms single-arranged particles in the middle, and the particles are surrounded by the sheath flow at four sides;
of the 3 outflow ends, the middle is an outflow end (waste liquid channel) of the sample flow, and the two sides are collecting channels; the particles sorted by the optical tweezers are deflected by the optical tweezers and flow out through the collecting channel.
Further, the system of the invention also comprises a light source system, wherein the light source system comprises an LED light source and a laser light source; wherein:
the LED light source generates white LED light which is focused by the lens to be used as an indicating light source of particles, and the indicating light source irradiates from the upper part of the micro-fluidic chip particle sample injection system;
the laser light source generates near-infrared laser, the unpolarized near-infrared laser is decomposed into two beams of different polarized light including P light and S light by the adjustable polarization type attenuator, the energy ratio of the two beams of laser is adjusted at will, and the two beams of polarized laser irradiate from the lower part of the micro-fluidic chip particle sample feeding system.
Furthermore, in the two beams of polarized laser, the S light is modulated by the acousto-optic modulator, and the acousto-optic modulator is used for modulating the intensity and the deflection angle of the laser simultaneously; the focusing light spots of the P light and the S light are respectively positioned at the upstream and the downstream of the microfluidic chip and are respectively used as an excitation light source and a capture light source; the S light passes through the beam expanding system and is focused by the objective lens to form an optical trap, and the optical trap is used for trapping particles.
Further, the wavelength of the near-infrared laser of the present invention is 808nm, 980nm, or 1064 nm.
Furthermore, the signal detection system of the invention comprises a plurality of photomultiplier tubes PMT and an InGaAs detector, which are respectively used for detecting the up-conversion luminescence signal and the forward scattering light signal of the particles, and the number of detection channels of the up-conversion luminescence signal is increased according to the requirement to construct a multi-channel detection system.
The invention provides a multifunctional optical tweezers method based on a microfluidic chip, which comprises the following steps:
the method comprises the following steps: coupling an up-conversion fluorescent probe to the surface of the particles, injecting the formed single-particle suspension into a channel of a micro-fluidic chip particle sample injection system, and enabling the sample flow to pass through a detection counting area and a capture sorting area in a single row according to a flow focusing principle;
step two: starting a near-infrared laser, and decomposing unpolarized near-infrared laser into two beams of different polarized light including P light and S light by the laser focused by an objective lens through an adjustable polarization type attenuator; the positions of the P light and the S light are observed and adjusted through a camera or a video camera, the power of the laser is adjusted, the P light acts on particles at the upstream in the channel to excite an up-conversion luminescence signal, and the S light forms a light trap at the downstream;
step three: and indicating the acousto-optic modulator to open or close the optical tweezers sorting system according to the strength of the up-conversion luminescence signal and the forward scattering light signal, so that the target particles enter the collecting channel under the action of the optical trapping force, and the particles which are not under the action of the optical trapping force enter the waste liquid channel, thereby carrying out real-time counting, quantitative analysis and sorting on the target particles.
Further, the probe of the present invention is an up-conversion nanomaterial.
Further, the types of target microparticles to be detected according to the present invention include: cells, cell exosomes, and nucleic acids, proteins, viruses, small molecules and metal ions enriched with microspheres.
Further, the type of target microparticle to be sorted according to the present invention is a microparticle, including a cell or a microsphere.
The invention has the following beneficial effects: the multifunctional optical tweezers system and the method based on the microfluidic chip have the following advantages:
1. the invention combines a microfluidic chip laboratory, an up-conversion material labeled biological sample and a multifunctional optical tweezers system, and provides a novel analysis and detection device which can analyze and separate various objects to be detected in real time.
2. The invention takes the up-conversion luminescent nano material with small particle size as the marking probe, and has small damage to the biological sample.
3. The multifunctional optical tweezers system constructed by the invention can detect and sort samples at the same time, and has high detection flux and sorting accuracy.
4. The invention divides the single laser into P light and S light, avoids using double lasers, is easy to respectively adjust the intensity of the two beams of light, simplifies the device and reduces the cost.
5. The invention uses a simple white light LED lighting source as an indicating light source of the sample at the same time.
6. The device of the invention adopts near infrared laser to excite the upconversion nanometer material to emit light, only a strong upconversion luminescent signal appears at a laser focusing focus, and the autofluorescence interference which is difficult to overcome in the conventional fluorescence detection method is overcome, so that when the method is used for analyzing a complex sample system, the anti-interference capability of the method is improved. In addition, forward scattered light signals generated by the action of laser and particles can accurately judge the occurrence of particles through the spectroscope. The combined analysis of the physical signal and the chemical signal is convenient for accurate subsequent sorting.
7. The device of the invention uses an InGaAs device to detect weak near-infrared scattered light signals, and uses a photomultiplier tube to detect up-conversion luminescence signals. Both detectors have the advantages of high signal-to-noise ratio, high sensitivity, high response speed and the like.
8. The device of the invention observes the visual field through a camera (video camera), and is convenient for quickly positioning the chip channel detection area.
9. When the invention is used for detecting biological samples, the invention has the characteristics of high flux, low background and high sensitivity.
Drawings
The invention will be further described with reference to the accompanying drawings and examples, in which:
FIG. 1 is a schematic view of the apparatus of the present invention;
FIG. 2 is a chip layout;
FIG. 3 schematic representation of tumor marker labeling up-converting nanomaterials;
FIG. 4 schematic representation of cell labeling up-conversion nanomaterials.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
In the specific embodiment of the invention, the multifunctional optical tweezers system for counting, detecting and sorting the microfluidic chip based on the upconversion luminescent nanomaterial labeled target combines the microfluidic chip laboratory and the upconversion luminescent nanomaterial labeled biological target to construct the multifunctional optical tweezers system, and deflects the captured laser by using the acousto-optic modulator, thereby realizing a system integrating counting, detecting and sorting, and being capable of being applied to the high-sensitivity detection of various biomolecules such as nucleic acid, protein or virus particles and other objects to be detected in a complex sample (such as whole serum and whole plasma), the sorting and diagnosis of tumor cells and the like.
In order to achieve the above purpose, the multifunctional optical tweezers system based on counting, detecting and sorting of the microfluidic chip comprises a microfluidic chip particle (cell, microsphere, etc.) sampling system, a forward scattering light counting system, an up-conversion luminescence signal detecting system and an optical tweezers deflection sorting system. The white light LED light source is focused by the lens and then is used as an indicating light source of the particles. Laser emitted by a near infrared laser vertically enters an adjustable polarization attenuator (VPBS), the laser is divided into two beams of different polarized light, reflected light is S light, transmitted light is P light, the P light enters a second adjustable polarization attenuator after passing through a lens group and a reflector, the reflected S light simultaneously enters the second adjustable polarization attenuator through an acousto-optic modulator (AOM) and the lens group, the two beams of light enter a 20-time objective rear pupil after passing through the reflector and a dichroic filter, the P light is focused by an objective, up-conversion luminescence excitation of particles is realized, the detection laser is called detection laser, the diameter of the beam is properly adjusted by the lens group, the diameter of a light spot focused in a micro channel by the objective is 15-20 microns, and the S light is focused by the objective to form a light trap for capturing sample particles. P light acts on the particles, forward scattered light signals are collected by a condenser lens, then are reflected by a dichroic filter and then enter a polarization spectroscope, then are detected by an InGaAs detector, up-conversion luminescent signals return and are detected by a Photomultiplier (PMT) through the dichroic filter, a signal detection system (DAQ data acquisition card) is combined with the up-conversion luminescent signals and the forward scattered light signals, whether the particles are target particles or not is judged while detection and counting are carried out, the optical tweezers deflect reaches a collection channel, the particles which are not acted by the action of optical trapping force enter a waste liquid channel, and therefore the system integrating counting, detection and sorting is achieved.
The non-polarized light is divided into two beams of polarized light by using the adjustable polarization type attenuator, so that the laser energy can be attenuated to the actually required power, and the scattered signal can be detected according to the polarized light. The adjustable polarization attenuator comprises two half-wave plates, an incident half-wave plate plays a role in adjusting the beam splitting ratio, and an emergent half-wave plate plays a role in adjusting the polarization direction of emergent light. If the polarization direction of the emergent side meets the requirement, the half-wave plate of the emergent side can be omitted. When a beam of unpolarized light enters perpendicularly, the beam splitter splits the beam into two beams of polarized light, the polarization states are perpendicular to each other, the reflected light is S light, and the transmitted light is P light. The energy ratio of the S light to the P light is related to the polarization direction of the incident light, so that the energy ratio of the P light to the S light can be changed by changing the polarization direction of the incident light.
The S light split by the spectroscope is modulated by an acousto-optic modulator, and the acousto-optic modulator can simultaneously modulate the intensity and the deflection angle of the laser.
The S light and the P light formed by splitting the same near-infrared laser beam have different effects. The focused spots of the P and S light are located upstream and downstream, respectively, in the microchannel and serve as excitation and capture light sources, respectively. The S light firstly enters the acousto-optic modulator, then is expanded by the lens group and is focused by the objective lens to form an optical trap.
The near-infrared laser is suitable for excitation of up-conversion luminescence signals, and the wavelength is 808nm, 980nm or 1064 nm.
The microfluidic chip channel design is mainly based on flowing and focusing a sample, so that sample particles pass through a detection area one by one.
The sample particles are marked by the up-conversion nano material, and the near-infrared laser is excited and detected, so that the light damage to the biological sample is small.
The detection and counting module detects the up-conversion luminescence by a photomultiplier tube and detects the forward scattered light by an InGaAs detector, and a CCD (or CMOS) camera (video camera) is used for imaging observation.
The acousto-optic modulator can modulate the intensity and direction of laser to form the actively controlled optical tweezers.
The signal detection system consists of a multi-channel data acquisition card and software.
The method for detecting and sorting the biological target by the multifunctional optical tweezers system based on counting, detecting and sorting of the microfluidic chip comprises the following specific steps:
coupling an up-conversion fluorescent probe to the surface of a particle, injecting a single-particle suspension into a micro-fluidic chip channel, and enabling a sample to pass through a preset detection area in a single row according to a flow focusing principle;
starting a near-infrared laser, adjusting the laser power of the laser focused by the objective lens, enabling the P light to act with the particles at the upstream, and ensuring to excite the up-conversion luminescence signal; the S light forms a light trap at the downstream;
and step three, indicating the acousto-optic modulator to open or close the optical tweezers according to the strength of the up-conversion luminescence signal and the forward scattering light signal, so that the target particles enter the collection channel, and the particles which are not acted by the optical trapping force enter the waste liquid channel, thereby carrying out real-time counting, quantitative analysis and sorting of the target particles.
The device of the invention uses a white light LED light source as an indicating light source of sample particles after being focused by a lens. Laser emitted by a semiconductor near-infrared fiber laser with the wavelength of 980nm vertically enters an adjustable polarization attenuator (VPBS) through a collimating lens and is divided into two beams of polarized light, reflected light is S light, transmitted light is P light, the P light enters a second adjustable polarization attenuator through a lens group and a reflecting mirror, meanwhile, the reflected S light enters the second adjustable polarization attenuator after being expanded through the lens group through an acousto-optic modulator, the two beams of light enter a 20 x objective rear pupil after passing through the reflecting mirror and a dichroic filter, the P light is used for up-conversion luminescence excitation of cells after being focused by an objective lens and is called detection laser, the diameter of the detection laser beam is properly adjusted through the lens group, the diameter of a light spot focused in a micro-channel by the objective lens is 15-20 mu m, and the S light is focused by the objective lens to form a light trap for capturing sample particles. The P light acts on the particle, the scattered light signal is collected by the condensing lens and then enters the polarization spectroscope after being reflected by the dichroic filter, then is detected by the InGaAs detector, the up-conversion luminescent signal then returns and is detected by Photomultiplier (PMT) through the dichroic filter, by signal detection system combination up-conversion luminescent signal and forward scattered light signal, judge whether the particle is the target when detecting and counting, the deflection of optical tweezers reaches the collection channel, the particle that does not receive the optical trapping power effect then gets into the waste liquid channel, thereby realize the count, detect and select separately in the system of an organic whole.
The invention adopts the up-conversion nano material to mark the object to be detected, and can adopt the up-conversion luminescent materials with different emission wavelengths to mark, thereby realizing the high-flux simultaneous detection of various particles. And the identification of the object to be detected is realized through the scattered light and the up-conversion luminescent signal.
The invention adopts a fluid dynamic focusing mode in the microfluidic chip to focus sample particles in a detection area of picoliter grade, and different sorting channels are designed to realize the separation of different objects to be detected.
The invention adopts the acousto-optic modulator AOM to realize the high flux sorting mode of active control.
The following examples are provided to further illustrate the detection and sorting methods of the present invention.
Example 1
Detection of tumor markers alpha-fetoprotein AFP and carcinoembryonic antigen CEA
The AFP and CEA antigens are respectively enriched by microspheres (3 microns and 5 microns) with different sizes of surface modified carboxyl groups through a double-site sandwich method, and the conversion material is used as a labeling material.
In the first step, immuno-encoded microspheres are prepared. Taking the microsphere with the surface carboxyl modified, activating the carboxyl on the surface of the microsphere through EDC/NHS reaction, and then respectively coupling AFP and CEA monoclonal antibodies at room temperature to obtain the immune microsphere.
Second, an upconversion probe is prepared. SMCC activates the surface amino-modified upconverting material followed by covalent coupling of another AFP and CEA monoclonal antibody.
And thirdly, enriching tumor markers respectively. The antigen capturing method by the double-site one-step method is characterized in that a certain amount of immune coding microsphere mixture with different sizes, an up-conversion nano probe and AFP and CEA antigens are respectively added into a centrifugal tube. And correspondingly enriching different antigens by using the immune microspheres with different sizes to obtain the coded microsphere compound.
And fourthly, injecting the coding microsphere compound into a microfluidic chip through an injection pump, enabling the coding microsphere compound to pass through a detection area in a single row under the action of sheath fluid entrained flow, exciting by near-infrared 980nm laser to generate an up-conversion luminescent signal and a scattered light signal, collecting the up-conversion luminescent signal by a Photomultiplier (PMT), and collecting a forward scattered light signal by an InGaAs (indium gallium arsenide) detector.
And fifthly, carrying out quantitative analysis. The method comprises the steps of measuring the strength of microspheres of a certain number of standard samples with different concentrations to prepare a standard curve, measuring a certain number of microspheres of unknown samples, and carrying out quantitative analysis by adopting a standard curve method.
Example 2
Sorting/typing of breast cancer cells
According to the different types of breast cancer cells (such as breast cancer cells MCF-7, SK-BR-3 and MDA-MB-453 of three different cell lines), green and red up-conversion luminescent nanomaterials are adopted to carry out double labeling on epithelial cell adhesion molecules (EpCAM) and human epidermal growth factor receptor-2 (HER2) on the surfaces of the breast cancer cells, antibodies and the up-conversion nanomaterials are coupled, then the antibodies and the up-conversion nanomaterials are specifically combined with antigens on target cells in a sample to be detected to form a target cell-antigen antibody-up-conversion particle compound, and then the compound is screened and enriched through optical tweezers. The method comprises the following specific steps:
the first step is as follows: an upconversion probe was prepared. SMCC activates two up-conversion nano materials with different emission wavelengths and surface amino modification, then two antibodies are respectively coupled covalently, and a target cell-antigen antibody-up-conversion nano particle compound is formed with a target cell in a sample to be detected.
The second step is that: injecting a sample to be detected into a micro-fluidic chip channel, enabling cells to pass through a detection area in a single row through flow focusing, enabling near-infrared 980nm laser to act with the cells to generate forward scattered light and up-conversion luminescent signals, enabling the scattered light signals to be used for counting, enabling the intensity of the two-channel luminescent signals to be used for judging the properties of the cells, sending an instruction after processing, and modulating captured laser through an acousto-optic modulator AOM so as to determine whether to open optical tweezers to deflect target cells to a sorting channel. At the instant of detecting the upconversion luminescent signal, the optical tweezers laser is in a closed state, when the green signal is significant (MCF-7), the optical tweezers capture the cell and move right in a short distance, and the laser is closed to release the cell, so that the cell is brought into a right channel; when the red signal is significant (SK-BR-3), the laser optical tweezers capture the cell and move left a short distance, while the laser is turned off to release the cell, with the result that the cell is brought into the left channel; when both the red and green signals are weak (MDA-MB-453), the cells continue to proceed straight into the middle channel.
Example 3
Detection and sorting of bacteria
According to the monoclonal antibody TEM-1 β -lactamase and the second antibody selective labeling anti-drug bacteria coupled with the upconversion nanometer material, the total bacteria are counted by combining with forward scattering light signals, and then the optical tweezers are utilized to sort and enrich, the specific steps are as follows:
bacteria are successively incubated with a monoclonal antibody TEM-1 β -lactamase and a covalent coupling secondary antibody green upconversion luminescent material.
The second step is that: similarly to the second step in example 2, the total number of bacteria was counted by forward scattered light, and whether the bacteria were drug-resistant was judged by up-conversion of the luminescence signal.
It will be understood that modifications and variations can be made by persons skilled in the art in light of the above teachings and all such modifications and variations are intended to be included within the scope of the invention as defined in the appended claims.
Claims (4)
1. A multifunctional optical tweezers system based on a microfluidic chip is characterized by comprising: the micro-fluidic chip particle sampling system, the counting system, the signal detection system and the optical tweezers sorting system; wherein:
the micro-fluidic chip particle sample introduction system is used for leading particles to pass through the micro-fluidic chip one by one along a certain path through hydrodynamic focusing, and a detection counting area and a capture sorting area are preset on the micro-fluidic chip; the unpolarized near-infrared laser is decomposed into two beams of polarized light including S light and P light by an adjustable polarization attenuator, and the energy ratio of the two beams of laser is adjusted arbitrarily; the acousto-optic modulator is used for modulating the intensity and the deflection angle of the S light at the same time; the focusing light spots of the P light and the S light are respectively positioned at the upstream and the downstream of the microfluidic chip and are respectively used as an excitation light source and a capture light source; the S light passes through a beam expanding system and is focused by an objective lens to form a light trap, and the light trap is used for capturing particles;
when the particles pass through the detection counting area, the signal detection system realizes simultaneous representation of multiple parameters of a luminescent signal and a forward scattering light signal of the particles by a weak light signal detection technology, realizes detection of the particles, and counts the number of the particles by the counting system;
when the particles pass through the capture sorting area, the optical tweezers sorting system deflects the particles of specific types according to the detection result of the particles, so that the sorting function of the particles under the action of the optical traps is realized; the signal detection system comprises a plurality of photomultiplier tubes PMT and an InGaAs detector, the PMT and the InGaAs detector are respectively used for detecting up-conversion luminescence signals and forward scattering light signals of particles, and the number of detection channels of the up-conversion luminescence signals is increased according to needs to construct a multi-channel detection system.
2. The multifunctional optical tweezers system based on the microfluidic chip as claimed in claim 1, wherein the microfluidic chip is provided with 3 inflow ends and 3 outflow ends, and the detection counting area and the capture sorting area are arranged at positions between the inflow ends and the outflow ends; wherein:
in the 3-way inflow end, sheath flow is arranged at two sides, and a sample flow of particles is arranged in the middle; the sample flow and the sheath flow simultaneously flow into the detection counting area and the capture sorting area, the sheath flows at two sides are used for ensuring that the sample flow forms single-arranged particles in the middle, and the particles are surrounded by the sheath flow at four sides;
the middle of the 3 outflow ends is the outflow end of a waste liquid channel of the sample flow, and the two sides are collecting channels; the particles sorted by the optical tweezers are deflected by the optical tweezers and flow out through the collecting channel.
3. A method of using the microfluidic chip based multifunctional optical tweezers system of claim 1, comprising the steps of:
the method comprises the following steps: coupling an up-conversion fluorescent probe to the surface of the particles, injecting the formed single-particle suspension into a channel of a micro-fluidic chip particle sample injection system, and enabling the sample flow to pass through a detection counting area and a capture sorting area in a single row according to a flow focusing principle;
step two: starting a near-infrared laser, and decomposing unpolarized near-infrared laser into two beams of different polarized light including P light and S light by the laser focused by an objective lens through an adjustable polarization type attenuator; the positions of the P light and the S light are observed and adjusted through a camera or a video camera, the power of the laser is adjusted, the P light acts on particles at the upstream in the channel to excite an up-conversion luminescence signal, and the S light forms a light trap at the downstream;
step three: the acousto-optic modulator is indicated to open or close the optical tweezers sorting system according to the strength of the up-conversion luminescence signal and the forward scattering light signal, so that target particles enter the collecting channel under the action of optical trapping force, and particles which are not under the action of optical trapping force enter the waste liquid channel, thereby carrying out real-time counting, quantitative analysis and sorting of the target particles; in quantitative analysis, the types of target particles detected include: cells, cell exosomes, and nucleic acids, proteins, viruses, small molecules and metal ions enriched with microspheres; the type of target particle to be sorted is a microparticle, including a cell or a microsphere.
4. The multifunctional optical tweezers system based on the microfluidic chip as claimed in claim 3, wherein the probe is an up-conversion nanomaterial.
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