Flow raman sorting method and high-luminous-flux flow raman sorting device

Abstract

A flow Raman sorting method and a high-luminous-flux flow Raman sorting device. The method comprises: performing coinciding focusing on a Raman laser (201) and a light-trap capture near-infrared laser (202), so as to form a Raman excitation detection site (204), and arranging the Raman excitation detection site (204) at a tip (112) of a terminal electrode (111) in a dielectric focusing electrode (110) of a dielectric microfluidic chip (100); focusing a photo-thermal oscillation near-infrared laser (203) on the terminal electrode (111) in the dielectric focusing electrode (110) of the dielectric microfluidic chip (100) without coinciding with the Raman excitation detection site (204); and by means of the switching between the light-trap capture near-infrared laser (202) and the photo-thermal oscillation near-infrared laser (203), generating a light-trap capture force by means of the light-trap capture near-infrared laser (202), so as to capture a single cell sample and fix the single cell sample at the Raman excitation detection site (204), or emitting bubbles by means of a photo-thermal effect generated by the photo-thermal oscillation near-infrared laser (203), such that cell samples adhering to the terminal electrode (111) in the dielectric focusing electrode (110) are oscillated and separated by means of the bubbles.

Description

Flow Raman sorting method and high-flux flow Raman sorting device

[0001] This application claims priority to the Chinese patent application filed with the China Patent Office on January 16, 2024, with application number 202410060771.3 and application name “A flow Raman sorting method and high-flux flow Raman sorting device”, the entire contents of which are incorporated by reference into this application. Technical Field

[0002] The present application belongs to the field of flow Raman sorting, and specifically relates to a flow Raman sorting method and a high-flux flow Raman sorting device. Background Art

[0003] The flow Raman sorter is an instrument for single-cell detection and sorting based on Raman spectroscopy technology. Since single cells are small, contain few components, and have a wide variety of types, their manipulation and analysis are extremely difficult. Therefore, how to keep single cells fixed in a flow state for Raman signal acquisition and how to collect high-sensitivity Raman spectra are the two most critical issues.

[0004] In terms of single cell fixation, there are existing implementations based on principles such as dielectric capture and optical trap capture. The two principles have different scopes of application, which limits the application of these two types of devices. In particular, in dielectric capture, it is very easy for a large number of cells to adhere to the electrode, so certain means are needed to oscillate and separate them to ensure that single cells can be detected and sorted.

[0005] Summary of the Invention

[0006] In a first aspect, the present application provides a flow Raman sorting method for use in a dielectric microfluidic chip, wherein the dielectric microfluidic chip includes a dielectric focusing electrode and a sorting electrode. The flow Raman sorting method includes:

[0007] Focusing the Raman laser and the optical trapping near-infrared laser together to form a Raman excitation detection site, and setting the Raman excitation detection site at the tip of the terminal electrode in the dielectric focusing electrode of the dielectric microfluidic chip;

[0008] Focusing the photothermal oscillation near-infrared laser on the end electrode of the dielectric focusing electrode of the dielectric microfluidic chip without overlapping with the Raman excitation detection site;

[0009] By switching between the light trap capture near-infrared laser and the photothermal oscillation near-infrared laser, the light trap capture near-infrared laser is used to generate a light trap capture force to capture and fix a single cell sample at the Raman excitation detection site, or bubbles are emitted through the photothermal effect generated by the photothermal oscillation near-infrared laser to enable the cell sample adhered to the end electrode in the dielectric focusing electrode to be oscillated and separated through the bubbles.

[0010] In some embodiments of the first aspect, a distance between a light spot formed by the photothermal oscillation near-infrared laser on the terminal electrode in the dielectric focusing electrode and the Raman excitation detection site is 5-15 μm.

[0011] In some embodiments of the first aspect, the light spot formed by the photothermal oscillation near-infrared laser on the terminal electrode in the dielectric focusing electrode and the Raman excitation detection site are both located on the tip center axis of the terminal electrode in the dielectric focusing electrode.

[0012] In a second aspect, the present application provides a near-infrared optical path system for a flow Raman sorting device, wherein the flow Raman sorting device sorts cells using a dielectric microfluidic chip, wherein the dielectric microfluidic chip includes a dielectric focusing electrode and a sorting electrode, and the near-infrared optical path system includes:

[0013] Near-infrared lasers;

[0014] a polarization beam splitter prism, configured to receive the near-infrared laser light emitted by the near-infrared laser and split the near-infrared laser light into a light trap captured near-infrared laser light and a photothermal oscillation near-infrared laser light;

[0015] A first shutter and a first Galilean beam expander are provided on the optical path of the photothermal oscillation near-infrared laser; a second shutter and a second Galilean beam expander are provided on the optical path of the light trap capturing the near-infrared laser;

[0016] An optical path adjustment component is used to receive the light trap capture near-infrared laser and the photothermal oscillation near-infrared laser and to adjust so that the Raman excitation detection site formed by the light trap capture near-infrared laser on the terminal electrode in the dielectric focusing electrode of the dielectric microfluidic chip and the light spot formed by the photothermal oscillation near-infrared laser optical path on the terminal electrode do not overlap, so that the light trap capture near-infrared laser acts on the tip of the terminal electrode to generate a light trap capture force, or the photothermal oscillation near-infrared laser acts on the terminal electrode to generate a bubble oscillation force through a photothermal effect.

[0017] In some embodiments of the second aspect, the optical path adjustment component includes:

[0018] a transflective beam displacement plate, which is arranged at an angle and coaxially arranged with the polarization beam splitter prism and the optical path of the photothermal oscillation near-infrared laser;

[0019] A first reflecting mirror and a second reflecting mirror are provided on an optical path of the light trap capturing the near-infrared laser; the first reflecting mirror is used to reflect the light trap capturing the near-infrared laser reflected by the polarization beam splitting prism; the second reflecting mirror is used to reflect the light trap capturing the near-infrared laser reflected by the first reflecting mirror onto the transflective beam displacement plate;

[0020] The third reflecting mirror is used to receive the light trap captured near-infrared laser reflected by the transflective beam displacement plate or the photothermal oscillation near-infrared laser transmitted by the transflective beam displacement plate.

[0021] An embodiment of the third aspect of the present application provides a high-throughput flow Raman sorting device, comprising:

[0022] A three-axis translation stage, wherein a visible light microscope objective lens for receiving the Raman signal collection and detection optical path and a near-infrared microscope objective lens for receiving the near-infrared laser emitted by the near-infrared optical path system are respectively provided on the upper and lower parts of the three-axis translation stage;

[0023] The Raman excitation optical path, the microscopic imaging optical path, and the Köhler coaxial illumination optical path are arranged through the Raman signal collection and detection optical path, and are respectively merged into the Raman signal collection and detection optical path through the first low-wavenumber Raman filter, the thin film spectroscope, and the flat plate spectroscope;

[0024] The thin film beam splitter and the flat plate beam splitter can be deflected to allow the microscopic imaging optical path and the Kohler coaxial illumination optical path to merge into the Raman signal collection and detection optical path or not to obstruct the Raman signal collection and detection optical path.

[0025] Another embodiment of the third aspect of the present application provides a high-flux flow Raman sorting device, including the near-infrared optical path system described in the second aspect above.

[0026] In some embodiments of the third aspect, the high-throughput flow Raman sorting apparatus further comprises:

[0027] A Raman signal acquisition and detection optical path system forms a Raman signal acquisition and detection optical path for acquiring and detecting Raman signals;

[0028] A Raman excitation optical path system, which forms a Raman excitation optical path for providing a laser source for Raman signal excitation, wherein the Raman excitation optical path is arranged along the Raman signal collection and detection optical path, and a first low-wavenumber Raman filter is arranged at the intersection of the Raman excitation optical path and the Raman signal collection and detection optical path, and the Raman excitation optical path is merged into the Raman signal collection and detection optical path through the first low-wavenumber Raman filter;

[0029] A microscopic imaging optical path system, which forms a microscopic imaging optical path for providing visualization of cell morphology and spatial position, wherein the microscopic imaging optical path is arranged along the Raman signal collection and detection optical path, and a thin film spectrometer is arranged near the intersection of the microscopic imaging optical path and the Raman signal collection and detection optical path;

[0030] A Köhler coaxial illumination optical path system, which forms a Köhler coaxial illumination optical path for providing illumination for microscopic imaging, wherein the Köhler coaxial illumination optical path is arranged along the Raman signal collection and detection optical path, and a flat plate beam splitter is arranged near the intersection of the Köhler coaxial illumination optical path and the Raman signal collection and detection optical path;

[0031] A three-axis translation stage, wherein a visible light microscope objective lens for receiving the Raman signal acquisition and detection optical path and a near-infrared microscope objective lens for receiving the near-infrared laser emitted by the near-infrared optical path system are respectively provided on the upper and lower parts of the three-axis translation stage;

[0032] The thin film beam splitter and the flat plate beam splitter can be deflected. By deflection, the thin film beam splitter and the flat plate beam splitter can enter the Raman signal collection and detection optical path and respectively reflect the microscopic imaging optical path and the Köhler coaxial illumination optical path into the visible light microscope objective to achieve microscopic imaging. Alternatively, the thin film beam splitter and the flat plate beam splitter can be reversely deflected to leave the Raman signal collection and detection optical path to avoid loss of light flux in the Raman signal collection and detection optical path.

[0033] In some embodiments of the third aspect, the Raman excitation optical path, the microscopic imaging optical path, and the Köhler coaxial illumination optical path are arranged in a parallel structure.

[0034] In some embodiments of the third aspect, the Raman signal acquisition and detection optical path system includes a conjugate mechanical spatial filtering component, an off-axis aspheric mirror, a low-wavenumber Raman filter b, a relay optical path component, and a spectrometer arranged in sequence along the Raman signal acquisition and detection optical path;

[0035] The Raman excitation optical path system includes a Raman single longitudinal mode laser, a Raman Galilean beam expansion component, and a conjugate Rayleigh line filter component which are sequentially arranged along the Raman excitation optical path;

[0036] The microscopic imaging optical system includes an imaging lens, a notch filter assembly, and a high-definition camera;

[0037] The Kohler coaxial illumination optical path system comprises an LED and a Kohler coaxial illumination component which are sequentially arranged along the Kohler coaxial illumination optical path.

[0038] A fourth aspect of the present application provides a sorting method based on the high-flux flow Raman sorting device described in any one of the above items, including a dielectric capture flow Raman sorting method and an optical trap capture flow Raman sorting method;

[0039] The dielectric capture flow Raman sorting method is:

[0040] The cell sample is loaded by air pressure drive, the thin film spectrometer and the flat plate spectrometer are rotated so that the microscopic imaging optical path and the Köhler coaxial illumination optical path are merged into the Raman signal collection and detection optical path to observe the cell state in real time, and a periodic dielectric signal is connected to the dielectric focusing electrode. The flow speed of the cell sample is controlled by controlling the dielectric on-off timing and adjusting the loading driving air pressure, so that the cell sample is focused through the dielectric focusing electrode to achieve a single cell focused flow and is captured at the tip of the terminal electrode in the dielectric focusing electrode, i.e., the Raman excitation detection site;

[0041] The Raman excitation light path is turned on to emit a Raman excitation source, and the cell sample at the tip of the terminal electrode in the dielectric focusing electrode is focused by the visible light microscope objective lens. The generated Raman signal enters the Raman signal collection and detection light path, and the thin film spectrometer and the flat plate spectrometer are rotated so that the rotating thin film spectrometer and the flat plate spectrometer leave the Raman signal collection and detection light path, thereby forming a single cell Raman spectrum to provide a criterion for sorting;

[0042] The host computer determines and identifies the cells, and selects the cells by connecting the selection electrodes to periodic dielectric signals;

[0043] In the above process, the optical path of the light trap capturing near-infrared laser is kept closed, the optical path of the photothermal oscillation near-infrared laser is opened at intervals, the position of the photothermal oscillation near-infrared laser is adjusted by adjusting the optical path adjustment component, and a bubble oscillation force is generated by the photothermal effect to oscillate and separate cells adhered to the tip of the terminal electrode in the dielectric focusing electrode;

[0044] The optical trap capture flow Raman sorting method is:

[0045] The cell sample is loaded by air pressure driving, the thin film beam splitter and the flat plate beam splitter are rotated so that the microscopic imaging optical path and the Köhler coaxial illumination optical path merge into the Raman signal acquisition and detection optical path to observe the cell state in real time, a double layer of sheath fluid is introduced into the dielectric microfluidic chip to form an intervening flow, and the loading driving air pressure is adjusted to control the sample flow rate and the sheath fluid flow rate, so that the cell sample is finally formed into a single cell assembly line in the central flow field of the Raman excitation detection site of the dielectric microfluidic chip;

[0046] Turning on the light trap to capture the near-infrared laser light path, capturing the cell sample at the Raman excitation detection site by the light trap to capture the near-infrared laser, and simultaneously suspending the sample loading driving air pressure;

[0047] The Raman excitation light path is turned on to emit a Raman excitation source, and the cell sample at the tip of the terminal electrode in the dielectric focusing electrode is focused by the visible light microscope objective lens. The generated Raman signal enters the Raman signal collection and detection light path, and the thin film spectrometer and the flat plate spectrometer are rotated so that the thin film spectrometer and the flat plate spectrometer leave the Raman signal collection and detection light path, thereby forming a single cell Raman spectrum to provide a criterion for sorting;

[0048] The host computer determines and identifies the cell samples captured by the optical trap and sorts them by moving the three-axis translation stage.

[0049] In some embodiments of the fourth aspect, the cell sample size applicable to the dielectric capture flow Raman sorting method is 5-60 μm; the cell sample size applicable to the optical trap capture flow Raman sorting method is less than 5 μm.

[0050] Compared with the prior art, the present invention has the following advantages:

[0051] 1. The flow Raman sorting method, near-infrared optical path system and flow Raman sorting device provided in at least one embodiment of the present application are combined with a dielectric microfluidic chip to set up a light trap to capture near-infrared laser and a photothermal oscillation near-infrared laser, so as to capture the near-infrared laser through the light trap to generate a light trap capture force to capture a single cell sample and fix it at the Raman excitation detection site, or to emit bubbles through the photothermal effect generated by the photothermal oscillation near-infrared laser, so that the cell sample adhered to the end electrode in the dielectric focusing electrode is oscillated and separated by the bubbles, thereby realizing the organic combination of light trap capture and dielectric capture (including sample oscillation function), which not only greatly reduces the instrument cost, but also effectively broadens the application range of the instrument and has strong versatility.

[0052] 2. At least one embodiment of the present application combines light trap capture and dielectric capture (including sample oscillation function) to design a near-infrared optical path. Through a near-infrared laser, the organic combination of light trap capture and dielectric capture (including sample oscillation function) can be simultaneously realized. The switching is convenient, flexible, fast, simple and reliable during use.

[0053] 3. The flow Raman sorting device provided in at least one embodiment of the present application is designed with an optical path structure based on common optical path switching. On the one hand, it can efficiently and losslessly irradiate the sample with the excitation light; on the other hand, it can return the returned Raman signal to the Raman signal acquisition and detection optical path without interference and with the highest possible optical flux. The number of optical elements passed through during the Raman signal acquisition and detection process is minimized to the greatest extent, achieving low signal loss and short acquisition time, ultimately achieving the purpose of high-flux measurement. Experimental verification shows that the flow Raman sorting device provided by the present application, compared with commercial microscopic confocal Raman spectroscopy measurement systems, increases the Raman excitation light flux from the general 50% to 85%, and the Raman signal collection light flux from the general no more than 30% to 41.8%, a total increase of approximately 40% in collected light flux, enabling high-flux detection of living single cells.

[0054] 4. The flow Raman sorting device provided in at least one embodiment of the present application adopts a coaxial common optical path design and a planar layout for the overall optical path. Most optical components are placed perpendicular to the optical axis or at 45°, so that each optical path is relatively independent and easy to debug. The system integration is flexible and simple, which greatly reduces the system debugging and maintenance costs. The optical path layout has reasonable transitions, the overall curing degree of the device is high, the vibration and impact resistance is strong, and the stability is high. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] The drawings described herein are used to provide further understanding of the present application and constitute a part of the present application. The illustrative embodiments of the present application and their descriptions are used to explain the present application and do not constitute improper limitations on the present application.

[0056] FIG1 is a schematic diagram showing the distribution of optical trapping of near-infrared laser light and photothermal oscillation of near-infrared laser light in a dielectric microfluidic chip in this application;

[0057] FIG2 is a schematic diagram of the structure of a near-infrared optical path system provided in an embodiment of the present application;

[0058] FIG3 is a schematic diagram of a laser passing through a transflective beam displacement plate in the near-infrared optical path system of FIG2 ;

[0059] FIG4 is a schematic structural diagram of a Raman sorting device provided in an embodiment of the present application;

[0060] FIG5 is a schematic structural diagram of a Raman signal acquisition and detection optical path system in a Raman sorting device provided in an embodiment of the present application;

[0061] FIG6 is a schematic diagram of the structure of the Raman excitation optical system in the Raman sorting device provided in an embodiment of the present application;

[0062] FIG7 is a schematic structural diagram of a microscopic imaging optical system in a Raman sorting device provided in an embodiment of the present application;

[0063] FIG8 is a schematic structural diagram of a Köhler coaxial illumination optical path system in a Raman sorting device provided in an embodiment of the present application;

[0064] In the picture:

[0065] 100, dielectric microfluidic chip; 110, dielectric focusing electrode; 111, end electrode; 112, tip of end electrode; 120, sorting electrode;

[0066] 201. Raman laser; 202. Optical trapping of near-infrared laser; 203. Photothermal oscillation of near-infrared laser; 204. Raman excitation detection site; 205. Spot formed by photothermal oscillation of near-infrared laser on the terminal electrode;

[0067] 300, near-infrared optical path system; 310, near-infrared laser; 320, polarization beam splitter prism; 331, first shutter; 332, second shutter; 341, first Galilean beam expander; 342, second Galilean beam expander; 350, optical path adjustment assembly; 351, transflective beam displacement plate; 352, first reflector; 353, second reflector; 354, third reflector; 301, optical path of photothermal oscillation near-infrared laser; 302, optical path of near-infrared laser captured by light trap;

[0068] 400, Raman signal acquisition and detection optical path system; 410, conjugate mechanical spatial filter assembly; 420, off-axis aspheric mirror; 430, second low-wavenumber Raman filter; 440, relay optical path assembly; 450, spectrometer; 401, Raman signal acquisition and detection optical path;

[0069] 500, Raman excitation optical system; 510, Raman single longitudinal mode laser; 520, third Galilean beam expander; 530, conjugate Rayleigh line filter; 501, first low-wavenumber Raman filter; 502, Raman excitation optical path;

[0070] 600, microscopic imaging optical system; 610, high-definition camera; 620, notch filter assembly; 630, imaging lens; 601, thin film beam splitter; 602, microscopic imaging optical path;

[0071] 700, Köhler coaxial illumination optical path system; 710, LED; 720, Köhler coaxial illumination assembly; 701, plate beam splitter; 702, Köhler coaxial illumination optical path;

[0072] 801, three-axis translation stage; 802, visible light microscope objective lens; 803, near-infrared microscope objective lens;

[0073] 901. Serial network topology control component; 902. Host computer. DETAILED DESCRIPTION

[0074] In order to make the purpose, technical solutions and advantages of this application more clearly understood, the present application is described and illustrated below in conjunction with the accompanying drawings and examples. It should be understood that the specific embodiments described herein are merely used to explain this application and are not intended to limit this application. Based on the embodiments provided in this application, all other embodiments obtained by those of ordinary skill in the art without making any creative efforts are within the scope of protection of this application.

[0075] Obviously, the drawings described below are merely examples or embodiments of the present application. Those skilled in the art can, without inventive effort, apply the present application to other similar scenarios based on these drawings. Furthermore, it is also understood that, although the effort involved in such a development process may be complex and lengthy, for those skilled in the art related to the content disclosed in this application, changes in design, manufacturing, or production based on the technical content disclosed in this application are merely conventional technical means and should not be construed as an insufficiency of the content disclosed in this application.

[0076] References to "embodiments" in this application mean that a particular feature, structure, or characteristic described in connection with the embodiment may be included in at least one embodiment of the application. The appearance of this phrase in various places in the specification does not necessarily refer to the same embodiment, nor does it refer to independent or alternative embodiments that are mutually exclusive of other embodiments. It is understood, both explicitly and implicitly, by those skilled in the art that the embodiments described in this application may be combined with other embodiments unless there is a conflict.

[0077] In a first aspect, the present application provides a flow Raman sorting method for use in a dielectric microfluidic chip 100. As shown in FIG1 , the dielectric microfluidic chip 100 includes a dielectric focusing electrode 110 and a sorting electrode 120. The electrode adjacent to the sorting electrode 120 in the dielectric focusing electrode 110 is an end electrode 111. The end electrode 111 has a V-shaped structure, with the tip 112 of the V-shaped structure pointing toward the sorting electrode. The specific structural form of the dielectric focusing electrode 110 and the sorting electrode 120 is shown in FIG1 . Since the above structure is prior art, it will not be elaborated on in detail.

[0078] In some embodiments, as shown in FIG1 , the flow Raman sorting method includes: focusing a Raman laser 201 and a light trap capture near-infrared laser 202 to form a Raman excitation detection site 204, and setting the Raman excitation detection site 204 at the tip 112 of the end electrode 111 in the dielectric focusing electrode 110 of the dielectric microfluidic chip 100, that is, at the tip 112 of the end electrode 111 with a V-shaped structure at the rightmost end of the dielectric focusing electrode 110 in FIG1 ; focusing a photothermal oscillation near-infrared laser 203 on the dielectric microfluidic chip 100; 00 is on the terminal electrode 111 in the dielectric focusing electrode 110, and does not overlap with the Raman excitation detection site 204; by switching between the optical trap capture near-infrared laser 202 and the photothermal oscillation near-infrared laser 203, the optical trap capture force is generated by the optical trap capture near-infrared laser 202 to capture and fix the single cell sample at the Raman excitation detection site or the photothermal effect generated by the photothermal oscillation near-infrared laser 203 is used to emit bubbles, so that the cell sample adhered to the terminal electrode 111 in the dielectric focusing electrode 110 is oscillated and separated by the bubbles.

[0079] In the above embodiment, the optical trap capture near-infrared laser 202 and the photothermal oscillation near-infrared laser 203 are essentially both near-infrared lasers, but their focusing positions are different, and therefore the effects they produce are different; wherein, the optical trap capture near-infrared laser 202 is focused on the tip 112 of the terminal electrode 111, forming an optical trap, and a single cell entering the optical trap can be captured and then detected; the photothermal oscillation near-infrared laser 203 is focused on the terminal electrode 111, and a photothermal effect occurs between it and the terminal electrode 111, thereby generating bubbles, and under the action of the bubbles, the cells and the electrode are separated.

[0080] Since the end electrode 111 in the dielectric focusing electrode 110 of the dielectric microfluidic chip 100 not only needs to realize light trap capture but also needs to realize photothermal oscillation separation, it is possible to consider setting the width of the end electrode 111 in the dielectric focusing electrode 110 to be greater than the width of other electrodes in the dielectric focusing electrode 110 during setting.

[0081] In some embodiments, the spacing between the light spot 205 formed by the photothermal oscillation near-infrared laser 203 on the terminal electrode 111 in the dielectric focusing electrode 110 and the Raman excitation detection site 204 is 5-15 μm. Taking into account practical size issues and to avoid focusing the near-infrared laser on the same point, which can easily cause the heat energy caused by the photothermal effect to be unable to be quickly discharged within the small area of ​​the electrode, directly causing the electrode tip to be damaged by breakdown, a staggered and non-overlapping setting is adopted to reduce the probability of such problems. Preferably, the specific spacing is set to 8-12 μm.

[0082] In some embodiments, the light spot 205 formed by the photothermal oscillation near-infrared laser 203 on the terminal electrode 111 of the dielectric focusing electrode 110 and the Raman excitation detection site 204 are both located on the central axis of the tip of the terminal electrode 111 of the dielectric focusing electrode 110. During the actual sorting process, under the influence of dielectric force, cells will flow in the direction of the electrode tip. This arrangement can achieve a better photothermal oscillation effect. Here, the central axis of the terminal electrode tip refers to the axis passing through the tip of the terminal electrode and bisecting the angle between the tips of the V-shaped structure.

[0083] The second aspect of the present application provides a near-infrared optical path system 300 for a flow Raman sorting device, which can be used to implement the flow Raman sorting method provided by the first aspect above. The near-infrared optical path system 300 can provide corresponding optical trap capture near-infrared laser 202 and photothermal oscillation near-infrared laser 203.

[0084] In some embodiments, as shown in FIG2 , the near-infrared optical system 300 includes:

[0085] A near-infrared laser 310 is used as a laser source for light trapping and photothermal oscillation. Optionally, the wavelength of the laser light generated by the near-infrared laser 310 can be in the near-infrared band, such as 785 nm or 1064 nm. The selected near-infrared laser 310 can have the characteristics of high power (>500 mW), narrow linewidth (<1 MHz), and high stability (power stability <5%). The specific laser can be selected from existing lasers according to needs.

[0086] The polarization beam splitter prism 320 is used to receive the near-infrared laser light emitted by the near-infrared laser 310 and split it into the light trap captured near-infrared laser light 202 and the photothermal oscillation near-infrared laser light 203. Specifically, the near-infrared laser light is reflected by the polarization beam splitter prism 320 to form the light trap captured near-infrared laser light 202, and the near-infrared laser light is transmitted through the polarization beam splitter prism 320 to form the photothermal oscillation near-infrared laser light 203.

[0087] The first shutter 331 and the first Galileo beam expander 341 are arranged on the optical path 301 of the photothermal oscillation near-infrared laser, and the second shutter 332 and the second Galileo beam expander 342 are arranged on the optical path 302 of the light trap to capture the near-infrared laser; the shutters 331 and 332 are used to respectively control the switching of the opening and closing of the corresponding optical paths; the Galileo beam expanders 341 and 342 are used to collimate and expand the laser to a suitable aperture, for example, the beam expansion ratio is limited to 2:1-7:1 according to the different entrance pupils of the near-infrared microscope objective 803, and the size of the light spot 205 finally formed by the photothermal oscillation near-infrared laser 203 on the end electrode 111 in the dielectric focusing electrode 110 is adjusted as needed.

[0088] The optical path adjustment component 350 is used to receive the light trap capture near-infrared laser 202 and the photothermal oscillation near-infrared laser 203 and adjust the Raman excitation detection site 204 formed by the light trap capture near-infrared laser 202 on the end electrode 111 in the dielectric focusing electrode 110 of the dielectric microfluidic chip 100 and the light spot 205 formed by the photothermal oscillation near-infrared laser 203 on the end electrode 111 in the dielectric focusing electrode 110 of the dielectric microfluidic chip 100 to not overlap, so that the light trap capture near-infrared laser 202 acts on the tip 112 of the end electrode 111 in the dielectric focusing electrode 110 to generate a light trap capture force, or the photothermal oscillation near-infrared laser 203 acts on the end electrode 111 in the dielectric focusing electrode 110 to generate a bubble oscillation force through the photothermal effect. The optical path adjustment component 350 can ensure that the distance between the light spot 205 formed by the photothermal oscillation near-infrared laser 203 on the end electrode 111 in the dielectric focusing electrode 110 and the Raman excitation detection site 204 is within the range of 5-15 μm.

[0089] In some embodiments, as shown in FIG2 , the optical path adjustment component 350 includes:

[0090] The transflective beam displacement plate 351 is arranged at an angle and is coaxial with the polarization beam splitting prism 320 and the optical path 301 of the photothermal oscillation near-infrared laser. The transflective beam displacement plate 8 can transmit a portion of light and reflect another portion of light. The reflected light is displaced relative to the original optical axis due to refraction. The transflective beam displacement plate can specifically be a flat beam splitter. The transmission and reflection ratio of the flat beam splitter can be 30:70 to 70:30, for example, 30:70, 50:50, or 70:30, and can be selected according to actual needs. Here, the transflective beam displacement plate 351 is arranged at an angle relative to the optical axis of the photothermal oscillation near-infrared laser.

[0091] A first reflector 352 and a second reflector 353 are provided on the optical path 302 of the light trap capturing the near-infrared laser light; the first reflector 352 is used to reflect the light trap capturing the near-infrared laser light 202 reflected by the polarization beam splitting prism 320, and is optionally arranged at an angle of 45° relative to the optical axis; the second reflector 353 is used to reflect the light trap capturing the near-infrared laser light 202 reflected by the first reflector 352 onto the transflective beam displacement plate 351, and is optionally arranged at an angle of 45° relative to the optical axis;

[0092] The third reflector 354 is configured to receive the light-trap captured near-infrared laser light 202 reflected by the transflective beam displacement plate 351 or the photothermally oscillated near-infrared laser light 203 transmitted by the transflective beam displacement plate 351. The light-trap captured near-infrared laser light 202 and the photothermally oscillated near-infrared laser light 203 are reflected by the third reflector 354 and focused at different locations on the end electrode of the dielectric focusing electrode 110.

[0093] In the aforementioned optical path adjustment assembly, after the photothermally oscillating near-infrared laser light 203 passes through the transflective beam displacement plate 351, it undergoes a displacement relative to its original optical path due to refraction (see Figure 3). The magnitude of this displacement is proportional to the thickness of the transflective beam displacement plate 351. The optical path of the light-trap captured near-infrared laser light 202 after reflection from the transflective beam displacement plate 351 is spaced apart from and parallel to the optical path of the transmitted photothermally oscillating near-infrared laser light 203. When the transflective beam displacement plate 351 is tilted 135° to the optical axis and the third reflector 354 is tilted 45° to the optical axis, the light-trap captured near-infrared laser light 202 and the photothermally oscillating near-infrared laser light 203, after reflection from the third reflector 354, enter the objective lens in parallel, converging at a single point without forming a separation. Therefore, by adjusting the angles of the transflective beam displacement plate 351 and / or the third reflector 354, the focal positions of the light-trap captured near-infrared laser light 202 and the photothermally oscillating near-infrared laser light 203 on the terminal electrode 111 do not overlap. Specifically, the reflected beam displacement plate 351 and the third reflector 354 can be deflected to adjust the angle, so that the position of the light spot on the imaging screen can be adjusted during the sorting process and the spacing can be controlled within the range of 5-15 μm.

[0094] In the embodiment shown in FIG2 , the near-infrared laser light generated by the near-infrared laser 310 is split into two laser beams after passing through the polarization beam splitting prism 320. One laser beam is transmitted through the polarization beam splitting prism 320 to form a photothermal oscillation near-infrared laser light 203. The photothermal oscillation near-infrared laser light 203 passes through the first shutter 331 and the first Galilean beam expander 341 in sequence, is transmitted through the transflective beam displacement plate 351, and is reflected by the third reflector 354 into the near-infrared microscope objective 803. The other laser beam is reflected by the polarization beam splitting prism 320 to form a light trap to capture the near-infrared laser light 202. The light beam passes through the second shutter 332, the first reflector 352, the second Galilean beam expander 342, and the second reflector 353 in sequence to reach the transflective beam displacement plate 351, is reflected by the transflective beam displacement plate 351 to the third reflector 354, and then is reflected by the third reflector 354 into the near-infrared microscope objective 803.

[0095] It is understandable that in the scheme of the present application, all optical elements in the optical path are not exhaustively listed. For example, reflectors not described in the present application may be arranged on the optical path. These reflectors can be specifically arranged according to the direction of the optical path. This is common knowledge in the art, and it should not be considered that the disclosure of the present application is insufficient.

[0096] A third aspect of the present application provides a high-throughput flow Raman sorting device.

[0097] In some embodiments, as shown in FIG4 , the high-throughput flow Raman sorting device includes:

[0098] A three-axis translation stage 801 is provided with a visible light microscope objective lens 802 for receiving the Raman signal acquisition and detection optical path 401 and a near-infrared microscope objective lens 803 for receiving the near-infrared laser (the optical trap capture near-infrared laser 202 or the photothermal oscillation near-infrared laser 203) emitted by the above-mentioned near-infrared optical path system, respectively, above and below the three-axis translation stage 801;

[0099] The Raman excitation optical path 502, the microscopic imaging optical path 602, and the Köhler coaxial illumination optical path 702 are arranged through the Raman signal collection and detection optical path 401, and respectively pass through the first low-wavenumber Raman filter 501, the thin film beam splitter 601, and the flat plate beam splitter 701 to converge into the Raman signal collection and detection optical path 401;

[0100] The thin film beam splitter 601 and the flat plate beam splitter 701 can be deflected to allow the microscopic imaging optical path 602 and the Köhler coaxial illumination optical path 702 to merge into the Raman signal collection and detection optical path 401 or not to obstruct the Raman signal collection and detection optical path 401 .

[0101] In other embodiments, as shown in Figure 4, the high-throughput flow Raman sorting device includes: a near-infrared optical path system 300 provided according to any embodiment of the second aspect of the present application, to provide an optical path 302 for light trapping to capture near-infrared laser and an optical path 301 for photothermal oscillation of near-infrared laser.

[0102] The high-throughput Raman sorting device further comprises:

[0103] Raman signal acquisition and detection optical path system 400, which forms Raman signal acquisition and detection optical path 401, is used to acquire and detect Raman signals; Raman signal acquisition and detection optical path system 400 receives Raman signals emitted by visible light microscope objective lens 802 and transmits them to serial network topology control component 901 and host computer 902 for detection and analysis, forming Raman signal acquisition and detection optical path 401;

[0104] A Raman excitation optical path system 500 forms a Raman excitation optical path 502 for providing a laser source for Raman signal excitation. The Raman excitation optical path 502 is disposed through the Raman signal collection and detection optical path 401. A first low-wavenumber Raman filter 501 is disposed at the intersection of the Raman excitation optical path 502 and the Raman signal collection and detection optical path 401. The Raman excitation optical path 502 passes through the first low-wavenumber Raman filter 501 and merges into the Raman signal collection and detection optical path 401.

[0105] A microscopic imaging optical path system 600 forms a microscopic imaging optical path 602 for providing visualization of cell morphology and spatial position. The microscopic imaging optical path 602 is disposed along the Raman signal collection and detection optical path 401. The pellicle beam splitter 601 is disposed near the intersection of the microscopic imaging optical path 602 and the Raman signal collection and detection optical path 401.

[0106] A Köhler coaxial illumination optical path system 700 forms a Köhler coaxial illumination optical path 702 for providing illumination for microscopic imaging. The Köhler coaxial illumination optical path 702 is disposed along the Raman signal collection and detection optical path 401. A flat plate beam splitter 701 is disposed near the intersection of the Köhler coaxial illumination optical path 702 and the Raman signal collection and detection optical path 401.

[0107] A three-axis translation stage, with a visible light microscope objective lens 802 for receiving the Raman signal acquisition and detection optical path 401 and a near-infrared microscope objective lens 803 for receiving the near-infrared laser (the optical trap capture near-infrared laser 202 or the photothermal oscillation near-infrared laser 203) emitted by the near-infrared optical path system, respectively, disposed above and below the three-axis translation stage 801;

[0108] Among them, the thin film beam splitter 601 and the flat plate beam splitter 701 can be set to be deflected; by deflection, the thin film beam splitter 601 and the flat plate beam splitter 701 can enter the Raman signal collection and detection optical path 401 (the thin film beam splitter 601 and the flat plate beam splitter 701 are in the position shown by the solid line in Figure 4) and respectively reflect the microscopic imaging optical path 602 and the Köhler coaxial illumination optical path 702 into the visible light microscope objective 802 to achieve microscopic imaging; alternatively, by reversely deflecting the thin film beam splitter 601 and the flat plate beam splitter 701, they are made to leave the Raman signal collection and detection optical path 401 (the thin film beam splitter 601 and the flat plate beam splitter 701 are in the position shown by the dotted line in Figure 4) to avoid losing the light flux of the Raman signal collection and detection optical path 401.

[0109] By setting the thin film beam splitter 601 and the flat plate beam splitter 701 as a deflectable structure, when performing microscopic imaging, the thin film beam splitter 601 and the flat plate beam splitter 701 are deflected (for example, so that the two are respectively at a 45-degree state relative to the optical axes of the microscopic imaging light path 602 and the Kohler coaxial illumination light path 702, as shown by the solid line in FIG4 ), so that the microscopic imaging light path 602 and the Kohler coaxial illumination light path 702 are respectively merged into the Raman signal collection and detection light path through the thin film beam splitter 601 and the flat plate beam splitter 701. Microscopic imaging is performed in the optical path 401; when Raman signal collection and detection is performed, the thin film spectrometer 601 and the flat plate spectrometer 701 are deflected (for example, so that the two are at 90 degrees relative to the optical axes of the microscopic imaging optical path 602 and the Köhler coaxial illumination optical path 702, respectively, as shown by the dotted line in Figure 4), and the thin film spectrometer 601 and the flat plate spectrometer 701 are completely separated from the Raman signal collection and detection optical path 401, thereby eliminating the signal splitting loss due to the coaxial common optical path and performing high-throughput Raman signal collection. The first low-wavenumber Raman filter 501 is used for high-flux transmission Raman signals. The low-wavenumber Raman filter mentioned in this application refers to a filter with a wavenumber lower than 100cm -1 Raman filter.

[0110] The thin film beam splitter 601 and the flat plate beam splitter 701 are used for co-optical path switching, specifically, rotating the flat plate beam splitter 701 and the thin film beam splitter 601 simultaneously or non-simultaneously, for example, from a 45-degree position to a 90-degree position, so that the thin film beam splitter 601 and the flat plate beam splitter 701 are completely separated from the Raman signal collection and detection optical path 401, thereby eliminating the signal splitting loss due to the coaxial common optical path and performing high-throughput Raman signal collection. Optionally, the deflection of the thin film beam splitter 601 and the flat plate beam splitter 701 can be achieved by driving a turntable with an electric motor, so that the turntable can deflect the thin film beam splitter 601 and the flat plate beam splitter 701 at high speed (each switching is less than 1 second) and high repeatability (repeat positioning accuracy is less than 0.5μm). Specifically, those skilled in the art can achieve this by purchasing existing equipment or using existing means.

[0111] The visible light microscope objective lens 802 is used for microscopic imaging of the sample and excitation of the Raman signal; in some embodiments, a high-magnification (>50X), high NA (>0.8), ultra-flat, apochromatic visible light microscope objective lens is selected, for example, an ultra-flat apochromatic microscope objective lens. In addition, those skilled in the art can also select other microscope objective lenses according to actual conditions. In some embodiments, the three-axis translation stage 801 is made of a high-precision structure and is used to move the dielectric microfluidic chip 100 with the built-in sample. It can achieve movement in three directions. The chip can be precisely moved forward, backward, left, right, and up and down by DC motor control. The minimum step value of the movement is 20nm, the repeat positioning accuracy during the movement is 0.5μm, and the range of movement is ±20mm front and back, ±35mm left and right, and ±20mm up and down. Specifically, those skilled in the art can achieve this by purchasing existing equipment or using existing means.

[0112] In some embodiments, the Raman excitation optical path 502 , the microscopic imaging optical path 602 , and the Köhler coaxial illumination optical path 702 are arranged in parallel.

[0113] In some embodiments, as shown in FIG5 , a Raman signal acquisition and detection optical path system 400 includes a conjugate mechanical spatial filter assembly 410, an off-axis aspheric mirror 420, a second low-wavenumber Raman filter 430, a relay optical path assembly 440, and a spectrometer 450, sequentially arranged along a Raman signal acquisition and detection optical path 401. The Raman signal acquisition and detection optical path is the core optical path of the device, and its function is to acquire and detect Raman signals at high throughput. The conjugate mechanical spatial filter component 410 is used to achieve true confocal imaging of microscopic Raman, and can be specifically a conjugate mechanical spatial pinhole filter component, which is characterized in that the rotation of the pinhole disk in the component can be controlled by an optical encoding motor, and pinholes of different sizes can be rotated into the optical path as needed, thereby preventing unnecessary signals outside the focal plane of the microscope from reaching the detector, playing a role in spatial filtering, and improving the spatial resolution of the instrument (<1.5μm); the off-axis aspheric mirror 420 is used to collimate and deflect the Raman signal; the second low-wavenumber Raman filter 430 is used to further filter out the Rayleigh signal and the high-transmittance Raman signal. Optionally, the second low-wavenumber Raman filter 430 is selected to have ultra-high edge steepness (0.2%), ultra-low wavenumber (<50cm -1), a low-wavenumber Raman filter with high Rayleigh cutoff (OD>6), and high Raman signal transmittance (>93%); the relay optical path component 440 is used to image the Raman signal onto the slit of the spectrometer 450 with high NA (Numerical Aperture) matching; the reflector is used for optical path deflection; the spectrometer 450 is used to split and detect the Raman signal, and finally transmit it to the imaging workstation to form a single-cell Raman spectrum, thereby providing a criterion for sorting. It is understandable that the various components with the above-mentioned characteristics are only some optional embodiments of the present application. Those skilled in the art can achieve this by purchasing existing components that meet the requirements or using means in the prior art, and it should not be considered that the disclosure of this application is insufficient.

[0114] In some embodiments, as shown in FIG6 , the Raman excitation optical path system 500 includes a Raman single longitudinal mode laser 510, a third Galilean beam expander 520, and a conjugate Rayleigh line filter 530 sequentially arranged along the Raman excitation optical path 502. Specifically, the function of the Raman excitation optical path system 500 is to provide a narrow linewidth, highly stable Raman signal excitation source. The Raman single longitudinal mode laser 510 provides a laser source for exciting Raman signals, and its wavelength can be in the visible light band such as 514.5 nm, 532 nm, and 632.8 nm. Lasers with high power (>200 mW), narrow linewidth (<1 MHz), single longitudinal mode (M2 <1.1), and high stability (<2%) can be selected. These characteristics ensure that the laser spot has very high spatial resolution after microscopic imaging. resolution (<0.6μm); the third Galilean beam expander 520 is used to collimate and expand the laser to a suitable aperture. Optionally, its beam expansion ratio is limited to 2:1-7:1 according to the different entrance pupils of the visible light microscope objective 802; the conjugate Rayleigh line filter component 530 is used to filter out laser wavelengths other than Rayleigh light; the first low-wavenumber Raman filter 501 is used to highly reflect the excitation light and highly cut off the returned Rayleigh light, thereby highly transmitting the Raman signal. Specifically, a filter with high edge steepness (0.5%), low wavenumber (<100cm -1 ), a low-wavenumber Raman filter with high excitation light reflectivity (>94%) and high Raman signal transmittance (>93%). Corresponding reflectors can be set in the optical path for folding the optical path. It is understandable that the components with the above-mentioned characteristics are only some optional embodiments of the present application. Those skilled in the art can achieve them by purchasing existing components that meet the requirements or by means of existing technologies, and it should not be considered that the disclosure of this application is insufficient.

[0115] In some embodiments, as shown in FIG7 , the microscopic imaging optical path system 600 includes an imaging lens 630, a notch filter assembly 620, and a high-definition camera 610 sequentially arranged along the microscopic imaging optical path 602; the function of the microscopic imaging optical path system 600 is to provide visualization of cell morphology and spatial position, so as to achieve precise measurement of “what you see is what you get”, and an infinite microscopic imaging system can be used. The sample image is reflected coaxially and along the same optical path by the thin film beam splitter 601 onto the imaging lens 630, which is characterized by a small change in optical path (<5 μm), almost no ghosting, and no chromatic aberration in the focused light beam. The function of the imaging lens 630 is to focus the sample image onto the high-definition camera 610. In some embodiments, a wide-field imaging, achromatic, and diffraction-limited imaging lens can be selected. The function of the notch filter assembly 620 is to filter out Rayleigh light and near-infrared light to enable clear imaging and protect the photosensitive surface of the camera. In some embodiments, a notch filter assembly that provides high OD cutoff (OD>6) for Rayleigh light and near-infrared light can be selected. The high-definition camera 610 is used for sample imaging. In some embodiments, a camera with a large rake surface (≥1 inch) and high resolution (>12 million pixels) can be selected. Corresponding reflectors can be set in the optical path for folding the optical path. It is understood that the various components with the above-mentioned characteristics are only some optional embodiments of the present application. Those skilled in the art can achieve this by purchasing existing components that meet the requirements or using means in the prior art. It should not be considered that the disclosure of this application is insufficient.

[0116] In some embodiments, as shown in FIG8 , a Köhler coaxial illumination optical path system 700 includes an LED 710 and a Köhler coaxial illumination assembly 720 sequentially arranged along a Köhler coaxial illumination optical path 702. The Köhler coaxial illumination optical path system 700 provides suitable brightfield illumination for microscopic imaging. The LEDs utilize white light LEDs as the illumination source. The Köhler coaxial illumination assembly 720, designed based on the Köhler illumination principle, provides a highly uniform light field. The illumination source is coaxially directed into the visible light microscope objective 802 via a flat beam splitter 701.

[0117] In a fourth aspect, the present application provides a sorting method based on the above-mentioned high-flux flow Raman sorting device, including a dielectric capture flow Raman sorting method and an optical trap capture flow Raman sorting method;

[0118] The dielectric capture flow Raman sorting method is:

[0119] The cell sample is loaded by air pressure drive, and the thin film beam splitter 601 and the plate beam splitter 701 are rotated so that the microscopic imaging optical path 602 and the Köhler coaxial illumination optical path 702 merge into the Raman signal acquisition and detection optical path 401 to observe the cell state in real time. A periodic dielectric signal is connected to the dielectric focusing electrode 110, and the flow speed of the cell sample is controlled by controlling the dielectric on-off timing and adjusting the loading driving air pressure. The cell sample is focused through the dielectric focusing electrode 110 to achieve a focused flow of a single cell and is captured at the tip of the terminal electrode 111 in the dielectric focusing electrode 110, i.e., the Raman excitation detection site 204;

[0120] The Raman excitation light path 502 is turned on to emit a Raman excitation source, which is focused on the cell sample at the tip 112 of the terminal electrode 111 in the dielectric focusing electrode 110 through the visible light microscope objective 802. The generated Raman signal enters the Raman signal collection and detection light path 401 and passes through the rotating thin film beam splitter 601 and the flat plate beam splitter 701, causing the rotating thin film beam splitter 601 and the flat plate beam splitter 701 to leave the Raman signal collection and detection light path 401, thereby forming a single-cell Raman spectrum to provide a criterion for sorting.

[0121] The host computer 902 determines and identifies the cells and sorts the cells by connecting the periodic dielectric signal to the sorting electrode 120;

[0122] During the above process, the optical trap capture near-infrared laser path is kept closed, and the photothermal oscillation near-infrared laser path 301 is opened at intervals. The position of the photothermal oscillation near-infrared laser 203 is adjusted by adjusting the optical path adjustment component 350, and a bubble oscillation force is generated by the photothermal effect to oscillate and separate cells adhered to the tip 112 of the terminal electrode 111 in the dielectric focusing electrode 110.

[0123] The optical trap capture flow Raman sorting method is:

[0124] The cell sample is loaded by air pressure drive, and the thin film beam splitter 601 and the plate beam splitter 701 are rotated so that the microscopic imaging optical path 602 and the Köhler coaxial illumination optical path 702 merge into the Raman signal acquisition and detection optical path 401 to observe the cell state in real time. A double layer of sheath fluid is introduced into the dielectric microfluidic chip 100 to form an intercalation flow. The loading drive air pressure is adjusted to control the sample flow rate and the sheath fluid flow rate. Ultimately, the cell sample is passed through the central flow field of the Raman excitation detection site 204 on the dielectric microfluidic chip 100 to form a single cell pipeline.

[0125] The light trap capture near-infrared laser light path is turned on, and the cell sample at the Raman excitation detection site 204 is captured by the light trap capture near-infrared laser 202, and the sample driving pressure is paused at the same time;

[0126] The Raman excitation light path 502 is turned on to emit a Raman excitation source, which is focused on the cell sample at the tip 112 of the terminal electrode 111 in the dielectric focusing electrode 110 through the visible light microscope objective 802. The generated Raman signal enters the Raman signal collection and detection light path 401. The thin film beam splitter 601 and the flat plate beam splitter 701 are rotated so that the thin film beam splitter 601 and the flat plate beam splitter 701 leave the Raman signal collection and detection light path 401, thereby forming a single-cell Raman spectrum to provide a criterion for sorting.

[0127] The host computer 902 determines and identifies the cell sample captured by the optical trap by moving the three-axis translation stage 801 to sort the cell sample.

[0128] In some embodiments, the size of cell samples applicable to the dielectric capture flow Raman sorting method is 5-60 μm; the size of cell samples applicable to the optical trap capture flow Raman sorting method is less than 5 μm.

[0129] Although the present application has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein; and these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present application.

Claims

1. A flow Raman sorting method, which is used in a dielectric microfluidic chip, wherein the dielectric microfluidic chip comprises a dielectric focusing electrode and a sorting electrode, characterized in that: The flow Raman sorting method comprises: Focusing the Raman laser and the optical trap near-infrared laser to form a Raman excitation detection site, and setting the Raman excitation detection site at the tip of the terminal electrode in the dielectric focusing electrode of the dielectric microfluidic chip; Focusing the photothermal oscillation near-infrared laser on the end electrode in the dielectric focusing electrode of the dielectric microfluidic chip without overlapping with the Raman excitation detection site; By switching between the light trap capture near-infrared laser and the photothermal oscillation near-infrared laser, the light trap capture near-infrared laser can generate a light trap capture force to capture and fix a single cell sample at the Raman excitation detection site, or the photothermal effect generated by the photothermal oscillation near-infrared laser can emit bubbles to enable the cell sample adhered to the end electrode in the dielectric focusing electrode to be oscillated and separated through the bubbles.

2. The sorting method according to claim 1, characterized in that: The distance between the light spot formed by the photothermal oscillation near-infrared laser on the terminal electrode in the dielectric focusing electrode and the Raman excitation detection site is 5-15 μm.

3. The sorting method according to claim 1, characterized in that: The light spot formed by the photothermal oscillation near-infrared laser on the terminal electrode in the dielectric focusing electrode and the Raman excitation detection site are both located on the tip center axis of the terminal electrode in the dielectric focusing electrode.

4. A near-infrared optical path system for a flow Raman sorting device, wherein the flow Raman sorting device sorts cells by means of a dielectric microfluidic chip, wherein the dielectric microfluidic chip comprises a dielectric focusing electrode and a sorting electrode, wherein: The near-infrared optical path system comprises: Near-infrared lasers; A polarization beam splitter prism is used to receive the near-infrared laser light emitted by the near-infrared laser and split it into a light trap captured near-infrared laser light and a photothermal oscillation near-infrared laser light; A first shutter and a first Galilean beam expander are arranged on the optical path of the photothermal oscillation near-infrared laser; a second shutter and a second Galilean beam expander are arranged on the optical path of the light trap capturing the near-infrared laser; The optical path adjustment component is used to receive the light trap capture near-infrared laser and the photothermal oscillation near-infrared laser and adjust the Raman excitation detection site formed by the light trap capture near-infrared laser on the terminal electrode in the dielectric focusing electrode of the dielectric microfluidic chip and the light spot formed by the photothermal oscillation near-infrared laser optical path on the terminal electrode to not overlap, so that the light trap capture near-infrared laser acts on the tip of the terminal electrode to generate a light trap capture force, or the photothermal oscillation near-infrared laser is used to generate a light trap capture force. Near-infrared laser acts on the terminal electrode to generate bubble oscillation force through photothermal effect.

5. The near-infrared optical path system according to claim 4, characterized in that: The optical path adjustment component comprises: A transflective beam displacement plate is arranged in an inclined manner and is arranged coaxially with the polarization beam splitter prism and the optical path of the photothermal oscillation near-infrared laser; A first reflector and a second reflector are arranged on the optical path of the light trap capturing the near-infrared laser; the first reflector is used to reflect the light trap capturing the near-infrared laser reflected by the polarization beam splitter prism; the second reflector is used to reflect the light trap capturing the near-infrared laser reflected by the first reflector onto the transflective beam displacement plate; The third reflecting mirror is used to receive the light trap captured near-infrared laser reflected by the transflective beam displacement plate or the photothermal oscillation near-infrared laser transmitted by the transflective beam displacement plate.

6. A high flux flow Raman sorting device, characterized in that: include: A three-axis translation stage, wherein a visible light microscope objective lens for receiving a Raman signal collection and detection optical path and a near-infrared microscope objective lens for receiving a near-infrared laser emitted by the near-infrared optical path system as claimed in claim 4 or 5 are respectively provided at the upper and lower parts of the three-axis translation stage; The Raman excitation optical path, the microscopic imaging optical path, and the Kohler coaxial illumination optical path are arranged through the Raman signal collection and detection optical path, and are respectively merged into the Raman signal collection and detection optical path through the first low-wavenumber Raman filter, the thin-film spectroscope, and the flat-plate spectroscope; The thin film beam splitter and the flat plate beam splitter can be deflected, and the microscopic imaging optical path and the Kohler coaxial illumination optical path are merged into the Raman signal collection and detection optical path or do not hinder the Raman signal collection and detection optical path through deflection.

7. A high flux flow Raman sorting device, characterized in that: Comprising the near-infrared optical path system as described in claim 4 or 5.

8. The high flux flow Raman sorting device according to claim 7, characterized in that: Also includes: A Raman signal collection and detection optical path system, which forms a Raman signal collection and detection optical path for collecting and detecting Raman signals; A Raman excitation optical path system, which forms a Raman excitation optical path for providing a laser source for Raman signal excitation, wherein the Raman excitation optical path is arranged along the Raman signal collection and detection optical path, a first low-wavenumber Raman filter is arranged at the intersection of the Raman excitation optical path and the Raman signal collection and detection optical path, and the Raman excitation optical path is merged into the Raman signal collection and detection optical path through the first low-wavenumber Raman filter; A microscopic imaging optical path system forms a microscopic imaging optical path for providing visualized cell morphology and spatial position, wherein the microscopic imaging optical path is arranged along the Raman signal collection and detection optical path. The membrane spectroscope is arranged near the intersection of the microscopic imaging optical path and the Raman signal collection and detection optical path; A Kohler coaxial illumination optical path system, which forms a Kohler coaxial illumination optical path for providing illumination for microscopic imaging, wherein the Kohler coaxial illumination optical path is arranged along the Raman signal collection and detection optical path, and a flat beam splitter is arranged near the intersection of the Kohler coaxial illumination optical path and the Raman signal collection and detection optical path; A three-axis translation stage, wherein a visible light microscope objective lens for receiving the Raman signal collection and detection optical path and a near-infrared microscope objective lens for receiving the near-infrared laser emitted by the near-infrared optical path system are respectively provided at the upper and lower parts of the three-axis translation stage; Wherein, the thin film beam splitter and the flat plate beam splitter can be deflected; by deflection, the thin film beam splitter and the flat plate beam splitter can enter the Raman signal collection and detection optical path and respectively reflect the microscopic imaging optical path and the Kohler coaxial illumination optical path to the visible light microscope objective to achieve microscopic imaging; or, by reversely deflecting the thin film beam splitter and the flat plate beam splitter to leave the Raman signal collection and detection optical path, the loss of the light flux of the Raman signal collection and detection optical path can be avoided.

9. The high flux flow Raman sorting device according to claim 6 or 8, characterized in that: The Raman excitation optical path, the microscopic imaging optical path and the Kohler coaxial illumination optical path are arranged in a parallel structure.

10. The high flux flow Raman sorting device according to claim 6 or 8, characterized in that: The Raman signal collection and detection optical path system comprises a conjugate mechanical spatial filtering component, an off-axis aspheric mirror, a low-wavenumber Raman filter b, a relay optical path component, and a spectrometer which are sequentially arranged along the Raman signal collection and detection optical path; The Raman excitation optical path system comprises a Raman single longitudinal mode laser, a Raman Galilean beam expansion component, and a conjugate Rayleigh line filter component which are sequentially arranged along the Raman excitation optical path; The microscopic imaging optical path system includes an imaging lens, a notch filter assembly, and a high-definition camera; The Kohler coaxial illumination optical path system comprises LEDs and Kohler coaxial illumination components which are sequentially arranged along the Kohler coaxial illumination optical path.

11. A sorting method based on the high flux flow Raman sorting device according to any one of claims 6 to 10, characterized in that: Including dielectric capture flow Raman sorting method and optical trap capture flow Raman sorting method; The dielectric capture flow Raman sorting method is: The cell sample is loaded by air pressure driving, the thin film spectroscope and the flat plate spectroscope are rotated, the microscopic imaging optical path and the Kohler coaxial illumination optical path are merged into the Raman signal collection and detection optical path to observe the cell state in real time, and the dielectric focusing electrode is connected to a periodic dielectric An electrical signal controls the flow rate of the cell sample by controlling the dielectric on-off timing and adjusting the sample loading driving gas pressure, so that the cell sample is focused through the dielectric focusing electrode to achieve a single cell focusing flow and is captured at the tip of the terminal electrode in the dielectric focusing electrode, i.e., the Raman excitation detection site; The Raman excitation optical path is turned on to emit a Raman excitation source, and the cell sample at the tip of the terminal electrode in the dielectric focusing electrode is focused through the visible light microscope objective lens, and the generated Raman signal enters the Raman signal collection and detection optical path, and the thin film beam splitter and the flat plate beam splitter are rotated so that the rotating thin film beam splitter and the flat plate beam splitter leave the Raman signal collection and detection optical path, thereby forming a single cell Raman spectrum to provide a criterion for sorting; The host computer determines and identifies the cells, and selects the cells by connecting the selection electrodes to periodic dielectric signals; In the above process, the optical path of the light trap capturing near-infrared laser is kept closed, the optical path of the photothermal oscillation near-infrared laser is opened at intervals, the position of the photothermal oscillation near-infrared laser is adjusted by adjusting the optical path adjustment component, and a bubble oscillation force is generated by the photothermal effect to oscillate and separate the cells adhered to the tip of the terminal electrode in the dielectric focusing electrode; The optical trap capture flow Raman sorting method is: The cell sample is loaded by air pressure driving, the thin film beam splitter and the flat plate beam splitter are rotated so that the microscopic imaging optical path and the Kohler coaxial illumination optical path are merged into the Raman signal collection and detection optical path to observe the cell state in real time, a double-layer sheath fluid is introduced into the dielectric microfluidic chip to form an inclusion flow, and the loading driving air pressure is adjusted to control the sample flow rate and the sheath fluid flow rate, so that the cell sample finally forms a single cell assembly line in the central flow field of the Raman excitation detection site of the dielectric microfluidic chip; Turning on the light trap to capture the near-infrared laser light path, capturing the cell sample at the Raman excitation detection site by the light trap to capture the near-infrared laser, and suspending the sample loading driving air pressure at the same time; The Raman excitation optical path is turned on to emit a Raman excitation source, and the cell sample at the tip of the terminal electrode in the dielectric focusing electrode is focused by the visible light microscope objective lens, and the generated Raman signal enters the Raman signal collection and detection optical path, and the thin film spectroscope and the flat plate spectroscope are rotated so that the thin film spectroscope and the flat plate spectroscope leave the Raman signal collection and detection optical path, thereby forming a single cell Raman spectrum to provide a criterion for sorting; The host computer determines and identifies the cell samples captured by the optical trap by moving the three-axis translation stage.

12. The sorting method according to claim 11, characterized in that: The size of the cell sample applicable to the dielectric capture flow Raman sorting method is 5-60 μm; the size of the cell sample applicable to the optical trap capture flow Raman sorting method is less than 5 μm.

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