A cell sorting and SERS detection device based on surface acoustic standing wave

Abstract

This invention discloses a cell sorting and SERS detection device based on surface acoustic waves (SAWs), belonging to the field of microfluidics. The device includes a SAW generation module and a microfluidic channel structure module bonded to it. The SAW generation module includes a piezoelectric substrate, a first-stage interdigital transducer, and a second-stage interdigital transducer. The microfluidic channel structure module includes a channel substrate, a first-stage sorting module, a SERS detection module, and a second-stage sorting module. A function signal generator is connected to the interdigital transducers via wires, and a corresponding radio frequency signal is input to generate SAWs. The radiation force of the SAWs acts on the microfluidic channel for first-stage sorting of cells of different sizes. SERS signals are qualitatively detected in the cells after first-stage sorting by spectral comparison, and the second-stage interdigital transducers are switched on and off for secondary sorting. This invention, by comprehensively utilizing SAW and SERS technologies, can quickly and effectively perform qualitative detection and sorting of cells of different sizes.

Description

Technical Field

[0001] This invention relates to microfluidics, and in particular discloses a cell sorting and SERS detection device based on surface acoustic waves, belonging to the technical field of MEMS devices. Background Technology

[0002] With the rapid development of microfluidic chip technology, the manipulation of particles within microfluidics has become increasingly sophisticated. Precise manipulation of fluids and particles immersed in microsystems is a key requirement for microfluidics research. Currently, particles in microfluidic systems can be manipulated using various methods, such as acoustics, optics, electromagnetics, and fluid dynamics. Surface acoustic wave (SAW)-based particle sorting methods have shown considerable application potential due to their scale-free nature and good biocompatibility.

[0003] Acoustic-based intrafluid particle separation technology combines acoustic flow and acoustic radiant force (ARF) to continuously manipulate particles of various sizes. A pair of interdigital electrodes are photolithographically etched onto the surface of a lithium niobate piezoelectric material substrate. When a corresponding radio frequency signal is applied to the interdigital electrodes, surface acoustic waves (SAWs) are generated on the lithium niobate substrate surface. SAWs consist of traveling waves and standing surface acoustic waves (SSAWs). Through the vibration of the substrate surface, acoustic energy is coupled into the liquid, allowing for precise manipulation of the fluid and particles. SAWs are advantageous in microfluidic systems because they can manipulate micron-sized materials at high frequencies, causing less damage to cells compared to methods such as magnetostriction and centrifugation, thus better preserving cell integrity.

[0004] Surface-enhanced Raman scattering (SERS) is an optical phenomenon that significantly enhances the Raman signal of analytes on rough nanostructures under laser excitation. SERS-based detection technologies show potential for single-molecule detection with high sensitivity and specificity. Existing SERS microfluidic cell separation and detection chips mainly employ microfiltration and biocapture methods to immobilize cells and detect biomolecules on the cell surface using SERS nanoprobes. Microfiltration, by designing microstructures to filter larger cells, such as tumor cells in blood, can cause severe cell deformation, affecting cell viability. Biocapture methods, using antibodies, aptamers, and other biomolecules to recognize cells, immobilize cells on the channel surface, making cell reculturing and detection difficult. Furthermore, SERS nanoprobe-based cell detection methods also rely on biorecognition; the nanoprobes are difficult to remove from the cell surface, affecting subsequent cell reculturing and detection.

[0005] Therefore, what we need is a microfluidic device system that can separate cells of different particle sizes without damaging the cells, automatically sample them for SERS detection, and then separate them again. Summary of the Invention

[0006] The purpose of this invention is to address the shortcomings of the aforementioned background technology by providing a cell sorting and SERS detection device based on surface acoustic waves, solving the technical problem of long time from SERS preparation to sample detection, and achieving the invention objective of simultaneously performing SERS detection and sorting.

[0007] To achieve the above-mentioned objectives, the present invention employs the following technical solution:

[0008] A cell sorting and SERS detection device based on surface acoustic waves includes: a surface acoustic wave generation module and a microfluidic channel structure module bonded to the surface acoustic wave generation module.

[0009] The surface acoustic wave generation module includes: a piezoelectric substrate and a first interdigital transducer, a second interdigital transducer, a third interdigital transducer, and a fourth interdigital transducer printed on the piezoelectric substrate. The first interdigital transducer and the third interdigital transducer are symmetrically distributed to form a first-level interdigital transducer, and the second interdigital transducer and the fourth interdigital transducer are symmetrically distributed to form a second-level interdigital transducer.

[0010] The microfluidic channel structure module includes: a channel substrate and a primary sorting module, a SERS detection module, and a secondary sorting module fabricated on the channel substrate. The primary sorting module is located in the region where the primary interdigital transducer generates a standing wave field. Under the action of the standing wave field generated by the primary interdigital transducer, the primary sorting module performs primary sorting of cells of different particle sizes in the flowing mixed liquid. The SERS detection module is located in the region where the secondary interdigital transducer generates a standing wave field. The SERS detection module calibrates the target cells in the analyte and triggers the secondary interdigital transducer when the target cells flow through the SERS detection module. Under the action of the standing wave field generated by the secondary interdigital transducer, the analyte undergoes secondary sorting and flows into the secondary sorting module. The secondary sorting module separates the target cells from the analyte.

[0011] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the first interdigital transducer, the second interdigital transducer, the third interdigital transducer and the fourth interdigital transducer have the same structure, each including: a first electrode connection area, a second electrode connection area, and an interdigital electrode pair; the two electrodes in the interdigital electrode pair are respectively connected to the first electrode connection area and the second electrode connection area.

[0012] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the channel substrate is attached to the piezoelectric substrate, and the channel substrate is located in the region where the first-stage interdigital transducer generates the standing wave field and the region where the second-stage interdigital transducer generates the standing wave field. The first-stage sorting module is a first-stage sorting channel opened on the channel substrate, and the first-stage sorting channel has a first sheath flow inlet, a mixed sample inlet, a second sheath flow inlet, a non-detectable analyte output port and a detectable analyte output port. The mixed sample inlet is located between the first sheath flow inlet and the second sheath flow inlet.

[0013] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the SERS detection module is a SERS detection channel connected to the output port of the object to be detected, and the output port of the object to be detected serves as the inlet of the SERS detection channel.

[0014] As a further optimization of a cell sorting and SERS detection device based on surface acoustic waves, the secondary sorting module includes a secondary sorting channel connected to the SERS detection channel outlet, and a first collection outlet and a second collection outlet located at the end of the secondary sorting channel.

[0015] As a further optimization of a cell sorting and SERS detection device based on surface acoustic waves, the number of interdigital electrode pairs is 35~55, and the working wavelength of the interdigital electrode pairs is 160μm~240μm.

[0016] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the primary sorting channel is located on the central axis. The symmetry lines of the first and third interdigital transducers are inclined at an angle of 15° to 30° with the central axis of the primary sorting channel, and the symmetry lines of the second and fourth interdigital transducers are inclined at an angle of 15° to 30° with the central axis of the primary sorting channel.

[0017] As a further optimization of a cell sorting and SERS detection device based on surface acoustic waves, the width ratio of the first sheath flow inlet, the mixed sample inlet, and the second sheath flow inlet at the confluence point is 1:1:1. The mixed sample inlet and the primary sorting channel are on the same horizontal plane. The first sheath flow inlet and the primary sorting channel are inclined at an angle of 30° to 60°. The second sheath flow inlet and the primary sorting channel are inclined at an angle of 30° to 60°. The width ratio of the non-detectable material output port and the detectable material output port at the confluence point is 1:1. The angle between the non-detectable material output port and the central axis of the primary sorting channel is 45° to 60°. The angle between the detectable material output port and the central axis of the primary sorting channel is 45° to 60°.

[0018] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the SERS detection channel is a cavity with an inwardly concave inner wall structure. The narrowest width of the inwardly concave inner wall structure is 20~25μm, the radius of curvature of the inwardly concave inner wall structure is 1mm, and the cross section of the inwardly concave inner wall structure perpendicular to the fluid flow direction is one of trapezoidal, rectangular and square or any combination thereof. The SERS detection channel is formed by etching a silicon wafer and the inner wall is coated with gold nanoparticles with a diameter of 10 nm-20 nm.

[0019] As a further optimization of the cell sorting and SERS detection device based on surface acoustic waves, the height of the primary sorting channel is 120 μm, and the width of the primary sorting channel is equal to the width of the non-detection outlet and the width of the detection outlet. The width of the secondary sorting channel is the same as the width of the primary sorting channel.

[0020] The present invention, employing the above-described technical solution, has the following advantages compared to existing microfluidic chips based on SERS detection technology:

[0021] (1) The present invention effectively utilizes the surface deformation of the piezoelectric substrate to generate surface acoustic waves. The surface acoustic waves propagate and couple into the mixed liquid, which can precisely control the liquid and particles, generate different sizes of acoustic radiation force between cells of different particle sizes, thereby achieving one-time sorting of cells and realizing cell size sorting. Compared with conventional microfluidic cell capture chips, this non-contact cell manipulation will not damage the physicochemical properties and biological activity of cells.

[0022] (2) The present invention uses SERS spectroscopy to label target cells of the test material after size sorting. It can realize online label-free cell detection while sorting cells, and can quickly and efficiently analyze the molecular characteristics of cells while they pass through, thereby identifying different types of cells.

[0023] (3) The present invention can simultaneously achieve cell size sorting and molecular feature sorting in a single chip system, effectively enriching a small number of cells in complex biological samples and obtaining higher sorting purity. Attached Figure Description

[0024] Figure 1 This is a schematic diagram of the structure of the cell sorting and SERS detection device provided by the present invention.

[0025] Figure 2 This is a schematic diagram of the interdigital transducer in this invention.

[0026] Figure 3 This is a schematic diagram of the SERS detection channel in this invention.

[0027] Figure 4 This is a schematic diagram of the SERS detection channel of the device provided by the present invention.

[0028] Figure 5 This is a schematic diagram of the secondary detection area of ​​the device provided by the present invention for cells.

[0029] Figure 6 This is a schematic diagram of the fabrication process of the standing wave generation unit in the device provided by the present invention.

[0030] Figure 7 This is a three-dimensional structural diagram of the SERS detection channel in this invention.

[0031] The labels in the diagram are as follows: 1. Lithium niobate substrate; 2. First interdigital transducer; 3. Second interdigital transducer; 4. Channel substrate; 5. Primary sorting channel; 6. First sheath flow inlet; 7. Mixed sample inlet; 8. Second sheath flow inlet; 9. Non-detectable analyte outlet; 10. Detectable analyte outlet; 11. SERS detection channel; 12. First collection outlet; 13. Second collection outlet; 14. Third interdigital transducer; 15. Fourth interdigital transducer; 16. Interdigital electrode pair; 17. First electrode connection area; 18. Second electrode connection area. Detailed Implementation

[0032] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0033] Example 1: Cell sorting and SERS detection device based on surface acoustic waves

[0034] like Figure 1 As shown, a cell sorting and SERS detection device based on surface acoustic waves (SAWs) includes a SAW generation module and a microfluidic channel structure module. The SAW generation module includes a lithium niobate substrate 1 and a first interdigital transducer 2, a second interdigital transducer 3, a third interdigital transducer 14, and a fourth interdigital transducer 15 printed on the lithium niobate substrate 1. The first interdigital transducer 2 and the third interdigital transducer 14 are symmetrically distributed to form a first-stage interdigital transducer, and the second interdigital transducer 3 and the fourth interdigital transducer 15 are symmetrically distributed to form a second-stage interdigital transducer. The microfluidic channel structure module includes a channel substrate 4 and a first-stage sorting module, a SERS detection module, and a second-stage sorting module fabricated on the channel substrate. A surface acoustic wave (SAW) generation module and a microfluidic channel structure module were prepared separately. Then, the microfluidic channel structure module was placed above the SAW generation module, so that the first-stage sorting module was located in the region where the first-stage interdigital transducer generates the standing wave field, i.e., the first-stage sorting region, and the SERS detection module was located in the region where the second-stage interdigital transducer generates the standing wave field, i.e., the second-stage detection region. The two modules were then bonded together to obtain the above-mentioned device.

[0035] like Figure 1 As shown, the channel substrate 4 is made of polydimethylsiloxane (PDMS); the channel substrate 4 is attached to the lithium niobate substrate 1 and is located between the first-stage interdigital transducers and between the second-stage interdigital transducers.

[0036] like Figure 1 As shown, a primary sorting channel 5 is formed on the channel base 4. The primary sorting channel 5 is located on the central axis and has a first sheath flow inlet 6, a mixed sample inlet 7, a second sheath flow inlet 8, a non-detectable analyte outlet 9, and a analyte outlet 10. The mixed sample inlet 7 is located between the first sheath flow inlet 6 and the second sheath flow inlet 8. The primary sorting channel 5, its three inlets, and two outlets together form a primary sorting module. The first interdigital transducer 2 and the third interdigital transducer 14 are inclined relative to the primary sorting channel 5. The symmetry line of the first interdigital transducer 2 and the third interdigital transducer 14 forms an inclined angle with the central axis of the primary sorting channel 5. They are used to sort large-diameter cells in the mixed sample during the transmission of RF coupling signals through the interdigital electrodes in the primary interdigital transducer. The large-diameter cells, as analytes, flow into the SERS detection module through the analyte outlet 10, while the small-diameter cells in the mixed sample flow out from the non-detectable analyte outlet 9.

[0037] like Figure 1 As shown, the SERS detection module and the secondary sorting module are fabricated on the channel substrate 4. The SERS detection module is a SERS detection channel 11 connected to the output port 10 of the analyte. The output port 10 of the analyte serves as the inlet of the SERS detection channel 11. The secondary sorting module includes a secondary sorting channel connected to the outlet of the SERS detection channel 11 and a first collection outlet 12 and a second collection outlet 13 located at the end of the secondary sorting channel. The center position of the SERS detection channel 11 is placed under a Raman grating spectrometer for SERS signal detection. The secondary interdigital transducer is turned on or off by analyzing the SERS signal through the system, thereby controlling the opening and closing of the acoustic wave. That is, alignment and correction are performed through the SERS detection module. After being detected by the SERS detection module and sorted by the secondary sorting module, the analyte flows out through the first collection outlet 12 or the second collection outlet 13. This forms a surface acoustic wave (SAW) sorting device.

[0038] like Figure 2As shown, the interdigital transducer includes: a first electrode connection area 17, a second electrode connection area 18, and interdigital electrode pairs 16. The interdigital electrode pairs 16 are positioned between the first electrode connection area 17 and the second electrode connection area 18, with two electrodes in each pair alternately connected to the two electrode connection areas; that is, one electrode in the interdigital electrode pair is connected to the first electrode connection area 17, and the other electrode is connected to the second electrode connection area 18. In use, a signal source inputs an RF coupling signal to the second electrode connection area 18 connected to the interdigital electrode pair, while the first electrode connection area 17 is grounded. The RF coupling signal is obtained by superimposing two sinusoidal signals. The frequencies of these two sinusoidal signals are within the operating bandwidth of the two interdigital electrode pairs inputting the RF coupling signal. The symmetrically arranged first and third interdigital transducers generate a standing wave field, which separates the mixed samples flowing through the first-stage sorting channel 5 under the influence of the standing wave field. The operation of the second-stage interdigital transducer is the same as that of the first-stage interdigital transducer.

[0039] In this embodiment, the height of the primary sorting channel 5 is 120μm, the width of the primary sorting channel 5 is equal to the width of the non-detection output port 9 and the width of the detection output port 10, and the width of the primary sorting channel is the same as that of the secondary sorting channel and can be set to 300μm.

[0040] In this embodiment, the width ratio of the first sheath flow inlet 6, the mixed sample inlet 7, and the second sheath flow inlet 8 at the confluence point is 1:1:1. The width of the three inlets can be set to 150μm. The mixed sample inlet 7 is on the same horizontal plane as the primary sorting channel, and the first sheath flow inlet 6 and the second sheath flow inlet 8 on both sides are inclined at an angle of 30° to 60° with the primary sorting channel.

[0041] In this embodiment, the width ratio of the non-detection output port 9 and the detection output port 10 at the confluence point is 1:1. The width of the non-detection output port 9 and the detection output port 10 can be set to 300μm. The angle between the two output ports and the central axis of the primary sorting channel 5 is 45°~60°.

[0042] In this embodiment, the primary sorting area is inclined at an angle of 15° to 30° to the central axis of the primary sorting channel 5, that is, the line of symmetry between the first interdigital transducer 2 and the third interdigital transducer 14 is inclined at an angle of 15° to 30° to the central axis of the primary sorting channel 5; the angle between the central axis of the secondary sorting channel and the secondary detection area is 15° to 30°, that is, the line of symmetry between the second interdigital transducer 3 and the fourth interdigital transducer 15 is inclined at an angle of 15° to 30° to the central axis of the primary sorting channel 5.

[0043] In this embodiment, the number of interdigital electrode pairs n = 35~55 in each interdigital transducer; the operating wavelength of the interdigital electrode pairs is 160μm~240μm.

[0044] In this embodiment, as Figure 7 As shown, the SERS detection channel 11 is a cavity with an inwardly concave inner wall structure. The narrowest width of the inwardly concave inner wall structure is 20~25μm, and the radius of curvature of the inwardly concave inner wall structure is 1mm. The inner wall of the SERS detection channel 11 is coated with gold nanoparticles with a diameter of 10 nm-20 nm.

[0045] The specific method for sorting multi-size cells using the surface acoustic wave-based cell sorting and SERS detection device in this embodiment is as follows:

[0046] Step 1: Connect the first electrode connection area 17, which is connected to the first-stage interdigital transducer, to the ground wire, and input an RF coupling signal into the second electrode connection area 18, which is connected to the first-stage interdigital transducer, so that the interdigital electrode pairs receive sinusoidal signals within their own operating bandwidth evenly.

[0047] Step 2: Input the mixed liquid into the mixed sample inlet located in the middle, and input the sheath flow into the first sheath flow inlet 6 and the second sheath flow inlet 8 located on both sides; as the mixed liquid flows through the primary sorting area where the primary sorting channel is located, large-diameter cells in the sorted material are separated from the mixed fluid and enter the test material output port 10; finally, the test material output port 10 outputs large-diameter cells, and small-diameter cells are output from the non-test material output port 9;

[0048] Step 3: Before large-diameter cells enter SERS detection channel 11, the channel width only allows particles within the specified size to pass through. The particles are arranged in a straight line and pass through SERS detection channel 11 one by one, such as... Figure 3 , Figure 4 As shown;

[0049] Step 4: A laser is emitted into the SERS detection channel 11, which is connected to the output port 10 of the analyte. The laser excites molecular vibrations on the sample surface, generating SERS signals. Gold nanoparticles coated on the inner wall of the SERS detection channel 11 enhance the SERS signal. The sample-scattered SERS spectral signal is collected and recorded. The collected SERS spectral signal is compared with system data, i.e., the cell reference spectral library. The chemical composition of the sample is determined based on the spectral signal comparison results. Cells with matching spectra are recorded and a feedback signal is generated. The feedback signal is triggered when the recorded cell passes through the secondary detection area. An RF coupling signal is input to the interdigital electrode pair in the secondary interdigital transducer. The secondary interdigital transducer generates surface acoustic waves, separating the desired sample from cells of the same particle size. Figure 5 As shown.

[0050] The working principle of the cell sorting and SERS detection device based on surface acoustic waves in this embodiment is as follows: A first-stage interdigital transducer excites surface elastic waves using the inverse piezoelectric effect. These elastic waves propagate in the opposite direction and superimpose to form a standing wave field. The acoustic energy in the standing wave field enters the mixed liquid through Rayleigh angle refraction and is scattered on the surface of the particles, thus generating acoustic radiation force. The magnitude of the acoustic radiation force is proportional to the cube of the particle radius; the larger the particle radius, the greater the acoustic radiation force. Within the same time interval, particles with larger radii experience greater lateral displacement forces than particles with smaller radii. Therefore, the lateral displacement of larger particles is greater than that of smaller particles, allowing large-diameter cells to flow out from the detection output port 10 and small-diameter cells to flow out from the non-detection output port 9, thereby achieving primary cell sorting. The second-stage interdigital transducer, enabled by the SERS detection feedback signal, forms a standing wave field, separating the target sample from cells of the same particle size through the secondary sorting channel.

[0051] Example 2: Preparation method of a cell sorting and SERS detection device based on surface acoustic waves.

[0052] The cell sorting device proposed in this invention requires precise processing technology and must be manufactured in a cleanroom environment. The presence of large dust particles can cause defects in the manufactured device, leading to device failure.

[0053] like Figure 6 As shown, the fabrication method of the surface acoustic wave (SAW) generation module includes the following steps 1 to 5:

[0054] Step 1: Using lithium niobate (LiNbO3) crystal as a piezoelectric substrate, the piezoelectric substrate is ultrasonically cleaned with acetone solution, and then repeatedly cleaned with ethanol in an ultrasonic machine. The piezoelectric substrate is dried with nitrogen to obtain a ready-to-use lithium niobate substrate.

[0055] Step 2: Take the prepared lithium niobate substrate, apply a 3μm thick layer of photoresist to its surface using a 3000rpm spin coater, and then place it on a hot plate for pre-baking at 100℃ for 2 minutes.

[0056] Step 3: Place the photomask onto the lithium niobate substrate coated with photoresist, and use light with an intensity of 40 mJ / cm². 2 After ultraviolet light exposure, the mask is removed, and the lithium niobate substrate with the mask removed is placed in the developing solution for development.

[0057] Step 4: A 100 nm thick gold film is grown on the lithium niobate substrate by electron beam evaporation to form the first electrode connection region and the second electrode connection region.

[0058] Step 5: After the lithium niobate substrate with gold film is plated, it is inverted and immersed in acetone solution for 10 minutes, then ultrasonically cleaned for 10 minutes. The lithium niobate substrate is then removed and ultrasonically treated with deionized water to form interdigitated electrode pairs.

[0059] The fabrication method of the microfluidic channel structure module includes the following steps one through seven:

[0060] Step 1: Mix the PDMS prepolymer and crosslinking agent in a ratio of 10:1 and stir until homogeneous;

[0061] Step 2: Place the mixed PDMS into a vacuum container and evacuate until air bubbles are removed;

[0062] Step 3: Pour the PDMS mixture into the prepared silicon mold;

[0063] Step 4: Place the mold into a vacuum container to remove air bubbles again;

[0064] Step 5: Place the mold in an oven or constant temperature box, set the temperature to 70 to 80 degrees Celsius, and cure for 2 to 4 hours to cure the PDMS.

[0065] Step 6: After the PDMS has cured, remove the PDMS block from the silicon mold and cut it into the required microfluidic chip shape with scissors or a blade to form a channel substrate.

[0066] Step 7: Using a drilling tool or other micromachining tool, holes are made in the PDMS chip to create a primary sorting channel, a first sheath flow inlet, a mixed sample inlet, a second sheath flow inlet, a non-detectable analyte output port and a detectable analyte output port, a SERS detection channel, a first collection outlet and a second collection outlet, thus obtaining the PDMS chip. Gold nanoparticles are coated on the inner wall of the SERS detection channel.

[0067] The specific process of coating the inner wall of the SERS detection channel with gold nanoparticles in step seven includes the following steps A to F:

[0068] Step A involves mixing a gold salt compound, such as chloroauric acid (HAuCl4), with a reducing agent, such as sodium hypophosphite (NaH2PO2), to trigger a reduction reaction. The gold ions produced in the reaction are gradually reduced to gold atoms, forming gold nanoparticles.

[0069] Step B involves adding a surfactant, such as hexadecyltrimethylammonium bromide (CTAB), to the reduction reaction solution to regulate the size and shape of the gold nanoparticles, thereby controlling the reaction kinetics and thermodynamic parameters. Surface modification can also be used to change the hydrophilicity or hydrophobicity of the nanoparticles.

[0070] Step C: The synthesized gold nanoparticles are separated from the reduction reaction solution by centrifugation.

[0071] Step D involves suspending the gold nanoparticles in water or an organic solvent to ensure their stability and dispersibility.

[0072] Step E: Use a brush to evenly coat the gold nanoparticle solution onto both sides of the SERS detection channel.

[0073] Step F: Place the PDMS chip in a dry, dust-free environment until the gold nanoparticles adhere to the inner wall of the SERS detection channel.

[0074] The method for bonding a surface acoustic wave generation module and a microfluidic channel structure module specifically includes steps b:

[0075] Step a: Bond the PDMS chip to the lithium niobate substrate to form a closed microfluidic system;

[0076] Step b: The hydrophilicity of the PDMS surface can be improved by methods such as oxygen plasma treatment, so as to improve its interaction with the liquid.

[0077] In this embodiment, the piezoelectric substrate in step 1 is specifically made of 128°YX-cut lithium niobate material.

[0078] In this embodiment, the first electrode connection region and the second electrode connection region prepared in step 4 are deposited by methods such as electron beam thermal evaporation or magnetron sputtering. The material can be one of molybdenum, gold, platinum, copper, aluminum, silver, titanium, tungsten, nickel or any combination thereof, with a thickness of 50-300 nm. The pattern is formed by processes such as plasma etching, lift-off, and wet etching, with a lateral width of 50-70 μm.

[0079] In this embodiment, the polydimethylsiloxane (PDMS) mixture obtained in step one is prepared by mixing a specially made PDMS liquid and a curing agent in a 10:1 ratio, followed by vacuuming and drying.

[0080] In this embodiment, the SERS detection channel is formed by etching a silicon wafer. The inwardly recessed structure of the inner wall, with a cross-section perpendicular to the fluid flow direction, can be one of trapezoidal, rectangular, and square or any combination thereof. The depth of the SERS detection channel cavity is 100-200μm, and the lateral width is 500-5000μm.

[0081] The above specific implementation methods and embodiments are specific support for the technical ideas proposed in this invention, and should not be used to limit the scope of protection of this invention. Any equivalent changes or modifications made based on the technical solutions of this invention in accordance with the technical ideas proposed in this invention shall still fall within the scope of protection of this invention.

Claims

1. A cell sorting and SERS detection device based on surface acoustic waves, characterized in that, include: A surface acoustic wave (SAW) generation module, and a microfluidic channel structure module bonded to the SAW generation module; The surface acoustic wave generation module includes: a piezoelectric substrate and a first interdigital transducer, a second interdigital transducer, a third interdigital transducer, and a fourth interdigital transducer printed on the piezoelectric substrate. The first interdigital transducer and the third interdigital transducer are symmetrically distributed to form a first-level interdigital transducer, and the second interdigital transducer and the fourth interdigital transducer are symmetrically distributed to form a second-level interdigital transducer. The microfluidic channel structure module includes: a channel substrate and a primary sorting module, a SERS detection module, and a secondary sorting module fabricated on the channel substrate. The primary sorting module is located within the region where the primary interdigital transducer generates a standing wave field. Under the action of the standing wave field generated by the primary interdigital transducer, the primary sorting module performs primary sorting of cells of different particle sizes in the flowing mixed liquid. The SERS detection module is located within the region where the secondary interdigital transducer generates a standing wave field. The SERS detection module calibrates the target cells in the analyte and triggers the secondary interdigital transducer when the target cells flow through the SERS detection module. Under the action of the standing wave field generated by the secondary interdigital transducer, the analyte undergoes secondary sorting and flows into the secondary sorting module. The secondary sorting module separates the target cells from the analyte. The channel substrate is attached to the piezoelectric substrate, and the channel substrate is located in the region where the first-stage interdigital transducer generates the standing wave field and the region where the second-stage interdigital transducer generates the standing wave field. The first-stage sorting module is a first-stage sorting channel formed on the channel substrate, and the first-stage sorting channel has a first sheath flow inlet, a mixed sample inlet, a second sheath flow inlet, a non-detectable analyte outlet, and a analyte outlet. The mixed sample inlet is located between the first sheath flow inlet and the second sheath flow inlet. The SERS detection module is a SERS detection channel connected to the analyte outlet, and the analyte outlet serves as the inlet of the SERS detection channel. The second-stage sorting module includes a second-stage sorting channel connected to the outlet of the SERS detection channel and a first collection outlet and a second collection outlet located at the end of the second-stage sorting channel.

2. The cell sorting and SERS detection device based on surface acoustic waves according to claim 1, characterized in that, The first, second, third, and fourth interdigital transducers have the same structure, each including: a first electrode connection area, a second electrode connection area, and an interdigital electrode pair; the two electrodes in the interdigital electrode pair are respectively connected to the first electrode connection area and the second electrode connection area.

3. The cell sorting and SERS detection device based on surface acoustic waves according to claim 2, characterized in that, The number of interdigital electrode pairs is 35 to 55, and the working wavelength of the interdigital electrode pairs is 160 μm to 240 μm.

4. The cell sorting and SERS detection device based on surface acoustic waves according to claim 1, characterized in that, The primary sorting channel is located on the central axis. The lines of symmetry of the first and third interdigital transducers are inclined at an angle of 15° to 30° with the central axis of the primary sorting channel. The lines of symmetry of the second and fourth interdigital transducers are also inclined at an angle of 15° to 30° with the central axis of the primary sorting channel.

5. The cell sorting and SERS detection device based on surface acoustic waves according to claim 1, characterized in that, The width ratio of the first sheath flow inlet, the mixed sample inlet, and the second sheath flow inlet at the confluence point is 1:1:

1. The mixed sample inlet and the primary sorting channel are on the same horizontal plane. The first sheath flow inlet and the primary sorting channel are inclined at an angle of 30° to 60°. The second sheath flow inlet and the primary sorting channel are inclined at an angle of 30° to 60°. The width ratio of the non-detection output port to the detection output port at the confluence point is 1:

1. The angle between the non-detection output port and the central axis of the primary sorting channel is 45°~60°, and the angle between the detection output port and the central axis of the primary sorting channel is 45°~60°.

6. The cell sorting and SERS detection device based on surface acoustic waves according to claim 1, characterized in that, The SERS detection channel is a cavity with an inwardly recessed inner wall structure. The narrowest width of the inwardly recessed inner wall structure is 20~25μm, the radius of curvature of the inwardly recessed inner wall structure is 1mm, and the cross-section of the inwardly recessed inner wall structure perpendicular to the fluid flow direction is one of trapezoidal, rectangular, and square or any combination thereof. The SERS detection channel is formed by etching a silicon wafer and the inner wall is coated with gold nanoparticles with a diameter of 10 nm-20 nm.

7. The cell sorting and SERS detection device based on surface acoustic waves according to claim 1, characterized in that, The height of the primary sorting channel is 120μm. The width of the primary sorting channel is equal to the width of the non-detection outlet and the width of the detection outlet. The width of the secondary sorting channel is the same as the width of the primary sorting channel.

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