Femtosecond laser processing-based novel gold-silicon SERS (Surface Enhanced Raman Scattering) substrate research and tumor cell detection application

Abstract

The invention discloses a preparation method of a gold-silicon SERS (Surface Enhanced Raman Scattering) substrate based on femtosecond laser processing and application of a micro-fluidic chip adopting the gold-silicon SERS substrate prepared based on femtosecond laser processing in capture and detection of cancer cells. The base preparation method comprises the following basic steps: S1, processing a micro-pit array on the surface of a silicon wafer through femtosecond laser processing; s2, electrochemically depositing a gold nano spine-shaped structure at the micro-pit array after femtosecond laser processing to prepare an SERS substrate; s3, testing the SERS performance of the substrate; and S4, incubating the substrate aptamer and capturing cancer cells. According to the gold-silicon SERS substrate based on femtosecond laser processing, SERS signals of an object to be detected can be remarkably improved, a large specific surface area is provided, and more detection sites are provided for molecules to be detected. Besides, the micro-pit array and the surface nano spine structure of the SERS substrate can effectively capture the cancer cells, a new method is provided for capturing and detecting the cancer cells, and the SERS substrate has great significance in early screening and diagnosis of human prostate cancer and analysis and research of heterogeneity of the cancer cells.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of micro-nano sensing technology, and in particular, the present application relates to a novel gold-silicon SERS substrate preparation method based on femtosecond laser processing and application. BACKGROUND

[0002] In 1928, Indian physicist C.V. Raman discovered that every substance has unique spectral characteristics, which can be used to identify specific molecules in a sample. However, due to the extremely weak Raman scattering signal (only 10 -6 ~ 10 -10 ), this phenomenon is difficult to use for in-depth study of molecular structure and properties. Until 1974, the emergence of surface-enhanced Raman scattering (SERS) technology broke this bottleneck. SERS technology is a highly sensitive, non-destructive detection method. This phenomenon was first discovered by accident by British scientist Fleischmann when studying pyridine molecules adsorbed on the surface of a rough silver electrode. He observed an abnormal Raman signal enhancement phenomenon. Subsequently, scholars such as Van Duyne and Creighton confirmed through systematic experimental verification and theoretical calculation that this signal enhancement was not simply due to the increase in electrode surface area, but a new physical enhancement mechanism. This major discovery laid the theoretical foundation for SERS technology and promoted its development into an important tool for modern spectral analysis.

[0003] Femtosecond laser processing is a kind of laser with extremely short pulse width, usually referring to laser with pulse duration in the range of femtosecond. Femtosecond laser processing has extremely high peak power, and can release a large amount of energy in a very short time. Therefore, femtosecond laser processing can ablate any material. When femtosecond laser processing is irradiated to the surface of the material, the laser pulse photons are absorbed by the material to form hot electrons, which collide with the material lattice to achieve material removal, thereby processing micro-nano structures on the material surface. Electrochemical deposition is an important micro-nano manufacturing technology in micro-nano structure processing. By controlling the electric field and chemical reactions in the electrolyte, metal or alloy materials can be precisely deposited on conductive substrates to form micro or nano scale structures.

[0004] Microfluidic chip technology, a scientific technique capable of manipulating fluids at the micrometer scale, has the potential to condense essential laboratory functions in biology, chemistry, and other fields onto a chip measuring less than a few square centimeters. Currently, most mainstream microfluidic chips consist of a network of microchannels, with controllable fluids flowing throughout the chip, enabling the various functions required for conventional chemical or biological experiments. Its fundamental characteristic and greatest advantage lies in the ingenious combination and large-scale integration of multiple microstructured units within a tiny, controllable platform to accomplish detection, analysis, and other functions.

[0005] In the prior art, most of the SERS substrate manufacturing processes are costly and require harsh processing conditions, and the SERS detection sensitivity is relatively insufficient.

[0006] In view of the above-mentioned deficiencies in the prior art, the present invention is proposed. Summary of the Invention

[0007] The purpose of the present invention is to solve the problems of complex processing conditions in the process of preparing traditional SERS substrates for capturing and detecting cancer cells, and provide a new gold-silicon SERS substrate preparation method based on femtosecond laser processing, as well as the application of microfluidic chips prepared based on femtosecond laser processing in the capture and detection of cancer cells.

[0008] To achieve the above object, the technical solution provided by the present invention is:

[0009] A method for preparing a gold-silicon SERS substrate based on femtosecond laser processing specifically comprises the following steps:

[0010] Step S1, femtosecond laser processing of silicon wafer;

[0011] Step S1.1: Select a prime-grade single-polish silicon wafer with a resistance of 1-10Ω·cm and a thickness of 500±10μm.

[0012] Step S1.2, cleaning the silicon wafer;

[0013] Step S1.3, design the laser path and write the G code;

[0014] Step S1.4, using femtosecond laser processing to process a micro-pit array on the surface of the silicon wafer;

[0015] Step S2, electrochemically depositing gold nano-spiky structures;

[0016] Step S2.1, cutting and cleaning the silicon wafer;

[0017] Step S2.2, preparing an electrolyte for electrochemical deposition;

[0018] Step S2.3, plasma-treating the cleaned silicon wafer for 150 seconds;

[0019] Step S2.4, electrochemically depositing gold nanospike structures in a two-electrode system;

[0020] The preparation of gold-silicon SERS substrate with micro-pit array on the surface is completed.

[0021] Step S3, SERS performance test of the substrate: after the substrate is subjected to oxygen plasma hydrophilic treatment, it is placed in MGITC solution and the SERS performance uniformity of the substrate is tested using a confocal Raman spectrometer;

[0022] Step S4, substrate aptamer incubation and capture of cancer cells;

[0023] The preferred technical solution provided by the present invention is:

[0024] The step S1.2 is specifically as follows:

[0025] The silicon wafer was ultrasonically cleaned in deionized water and anhydrous ethanol for 10 minutes respectively, and then dried with nitrogen gas after cleaning;

[0026] The preferred technical solution provided by the present invention is:

[0027] The step S1.3 is specifically as follows:

[0028] Step S1.3.1: Design a 100 x 100 micropit array, with a spacing of approximately 10 μm between each micropit and each micropit approximately 20 μm long, 20 μm wide, and 15 μm deep.

[0029] Step S1.3.2, write the G code according to step S1.3.1;

[0030] The preferred technical solution provided by the present invention is:

[0031] The step S1.4 is specifically as follows:

[0032] Step S1.4.1: Fix the silicon wafer on a three-dimensional high-precision mobile platform, adjust the laser optical path, and assemble the objective lens to 10X / 0.25;

[0033] Step S1.4.2: Turn on the CCD camera and focus on the silicon wafer plane until the image is clear;

[0034] Step S1.4.3: Once the image is clear, slightly move the Z and X / Y axes of the 3D high-precision mobile platform while simultaneously operating the optical switch to mark the laser focus. Observe the laser focus plane under an optical microscope. The Z-axis position corresponding to the point with the smallest spot size is the laser focus plane.

[0035] In step S1.4.4, the processing laser power is 10.0 W, the frequency is 50 kHz, the wavelength is 1030 nm, and the energy is 200 μJ. Using an attenuator and laser frequency division, the laser power is controlled to 0.150 W, the frequency is 5.0 kHz, and the energy is 30.0 μJ. G code is run to control the movement of the three-dimensional high-precision mobile platform while femtosecond laser processing is performed on the silicon wafer.

[0036] The preferred technical solution provided by the present invention is:

[0037] The step S2.1 is specifically as follows:

[0038] After femtosecond laser processing, the silicon wafers were cut and ultrasonically cleaned in deionized water, anhydrous ethanol, and acetone for 15 minutes respectively, and then dried with nitrogen gas after cleaning.

[0039] The preferred technical solution provided by the present invention is:

[0040] The step S2.2 is specifically as follows: preparing a 10 mM HAuCl4 solution and a 0.5 M HCl solution, adding equal volumes of the two solutions to the electrolytic cell, and then adding 1% Triton X-100 of the electrodeposition system to complete the electrolyte preparation;

[0041] The preferred technical solution provided by the present invention is:

[0042] The step S2.4 is specifically as follows:

[0043] Step S2.4.1: Use a silicon wafer as the cathode (working electrode) and a carbon rod as the anode (auxiliary electrode). Use a copper rod-platinum electrode clamp to secure and connect to an electrochemical workstation. Preheat the electrochemical workstation for 30 minutes before electrodeposition.

[0044] In step S2.4.2, connect the green clip WE to the working electrode and the red clip CE to the auxiliary electrode. Select i-tAmperometric i-tCurve chronoamperometry, set the potential to -3 V, the deposition time to 1800 s, the Sample Interval to 0.01 s, the Quiet Time to 0 s, and the Sensitivity to 1.e-003. Click OK to start electrodeposition.

[0045] Step S2.4.3: After the electrodeposition is completed, the sample is removed from the electrolytic cell and placed in an oven for drying;

[0046] The preferred technical solution provided by the present invention is:

[0047] The step S3 is specifically as follows:

[0048] The SERS substrate was treated with oxygen plasma for 90-120 seconds, then immersed in a 10-6 mol / L MGITC solution in the dark for 3-4 hours. Finally, it was removed and rinsed with deionized water to remove any free MGITC Raman reporter molecules not attached to the SERS substrate, and then placed in a petri dish to air-dry. The prepared SERS substrate was placed on the test sample stage of a Renishaw Raman spectrometer. Raman measurements were then performed on the SERS substrate, and the results were collected and statistically analyzed. The Renishaw Raman spectrometer test parameters were: 20× microscope objective, excitation light source: 633 nm, laser power: 1.7 mW to 5.5 mW, integration time: 1 s, integration count: 1, laser intensity: 5%. Data were collected and recorded from 10 random locations on the SERS substrate, and the average value was calculated and compared with the Raman signal of a standard gold-coated substrate.

[0049] The preferred technical solution provided by the present invention is:

[0050] The step S4 is specifically as follows:

[0051] Step S4.1, modifying the two specific aptamers on the SERS substrate; first, the prepared SERS substrate is hydrophilicized, then placed in a solution of wy-5a and PSMA aptamers, slowly shaken for 24 hours, and finally, the residual aptamers are rinsed with buffer to complete the substrate modification;

[0052] Step S4.2, add 30 μL of 2.5% glutaraldehyde solution to the substrate for fixation for 2-3 hours; glutaraldehyde is used to fix the cancer cells and maintain their morphology when they are captured;

[0053] Step S4.3, placing the SERS substrate in 30%, 50%, and 80% alcohol solutions for 2 min each;

[0054] Step S4.4: Place the SERS substrate on a sample stage with an inclination angle of 45 degrees and spray palladium for 120-150 seconds.

[0055] The cells captured on the SERS substrate can then be observed using SEM.

[0056] Wherein, the cancer cells refer to PC-3 tumor cells and LNCaP tumor cells in prostate cancer.

[0057] Compared with the prior art, the present invention has the following beneficial effects:

[0058] The present invention solves the problems of complex processing conditions and low SERS signal intensity in the preparation of traditional SERS substrates for capturing and detecting cancer cells, and provides a new idea, namely, integrating femtosecond laser processing to significantly increase the specific surface area of ​​the substrate and electrochemically deposited gold nanospike structures to provide more SERS hotspots to increase Raman scattering. The micro-pit array on the surface of the SERS substrate can effectively capture cancer cells, can be produced in large quantities, has high repeatability, is simple to prepare, and does not require processing with high-risk equipment. The prepared substrate has good Raman scattering enhancement effect, strong stability, is non-toxic and harmless to cells, and has little damage, and can be used for capturing and detecting cancer cells.

[0059] Explanation of terms:

[0060] PDMS – polydimethylsiloxane;

[0061] MGITC – Malachite green isothiocyanate solution. BRIEF DESCRIPTION OF THE DRAWINGS

[0062] Figure 1 Schematic diagram of the gold-silicon SERS substrate based on femtosecond laser processing and its application in capturing and detecting cancer cells of the present invention;

[0063] Figure 2 Schematic diagram of the preparation process of the gold-silicon SERS substrate based on femtosecond laser processing of the present invention;

[0064] Figure 3 is the SEM image of the prepared SERS substrate;

[0065] Figure 4 SEM image of cancer cells captured by substrate;

[0066] Figure 5 Raman signal detection of the prepared SERS substrate;

[0067] Figure 6 To detect the uniformity of the prepared SERS substrate. DETAILED DESCRIPTION

[0068] The technical solutions in the embodiments of the present application will be described clearly and completely below with reference to the accompanying drawings.

[0069] Figure 1 Schematic diagram of the overall structure of the gold-silicon SERS substrate based on femtosecond laser processing of the present invention; Figure 2 FIG. 1 is a schematic diagram of the preparation process of the gold-silicon SERS substrate based on femtosecond laser processing of the present invention; FIG. Figure 1 、 Figure 2 As shown, a method for preparing a gold-silicon SERS substrate based on femtosecond laser processing of the present invention comprises the following steps:

[0070] Step S1, femtosecond laser processing of silicon wafer;

[0071] Step S1.1: Select a prime-grade single-polish silicon wafer with a resistance of 1-10Ω.cm and a thickness of 500±10μm.

[0072] Step S1.2: ultrasonically clean the silicon wafer in deionized water and anhydrous ethanol for 10 minutes each, and blow dry with nitrogen gas after cleaning to remove residual organic solvents and small debris on the surface of the silicon wafer;

[0073] Step S1.3, design the laser path and write the G code;

[0074] The step S1.3 is specifically as follows:

[0075] Step S1.3.1: Design a 100 x 100 micropit array with a spacing of approximately 10 μm between each micropit. Each micropit is approximately 20 μm long, 20 μm wide, and 15 μm deep. The size of the micropit array is designed based on the size of the cancer cells.

[0076] Step S1.3.2: Write the G code according to step S1.3.1. The G code for femtosecond laser processing of a single micro-pit is: GDO OP8 S1 G00 X0.01 F0.5 G00 Y0.005 F0.5 G00 X-0.01F0.5 G00 Y0.005 F0.5 G00 X0.01 F0.5 G00 Y-0.01F0.5 GDO OP8 S0;

[0077] Step S1.4, using femtosecond laser processing to process a micro-pit array on the surface of the silicon wafer;

[0078] The step S1.4 is specifically as follows:

[0079] Step S1.4.1: Fix the silicon wafer on a three-dimensional high-precision mobile platform, adjust the laser optical path, and assemble the objective lens to 10X / 0.25;

[0080] Step S1.4.2: Turn on the CCD camera and focus on the silicon wafer plane until the image is clear;

[0081] Step S1.4.3: Once the image is clear, slightly move the Z and X / Y axes of the 3D high-precision mobile platform while simultaneously operating the optical switch to mark the laser focus. Observe the laser focus plane under an optical microscope. The Z-axis position corresponding to the point with the smallest spot size is the laser focus plane.

[0082] In step S1.4.4, the processing laser power is 10.0 W, the frequency is 50 kHz, the wavelength is 1030 nm, and the energy is 200 μJ. Using an attenuator and laser frequency division, the laser power is controlled to 0.150 W, the frequency is 5.0 kHz, and the energy is 30.0 μJ. G code is run to control the movement of the three-dimensional high-precision mobile platform while femtosecond laser processing is performed on the silicon wafer.

[0083] Step S2, electrochemically depositing gold nano-spike structures;

[0084] Step S2.1, cutting and cleaning the silicon wafer;

[0085] The step S2.1 is specifically as follows:

[0086] After femtosecond laser processing, the silicon wafers were cut and ultrasonically cleaned in deionized water, anhydrous ethanol, and acetone for 15 minutes respectively. After cleaning, they were blown dry with nitrogen to remove residual processing debris, fingerprints, and small debris on the surface of the silicon wafers.

[0087] Step S2.2, preparing an electrolyte for electrochemical deposition;

[0088] The step S2.2 is specifically as follows:

[0089] Prepare a 10mM HAuCl4 solution and a 0.5M HCl solution. Add equal volumes of the two solutions to the electrolytic cell. Then add 1% Triton X-100, which is the electrodeposition system. The purpose of adding Triton X-100 is to reduce the water tension of the electrolyte on the surface of the micropits, allowing the electrolyte to enter the bottom of the micropits. This completes the electrolyte preparation.

[0090] Step S2.3: Plasma-treating the cleaned silicon wafer for 150 seconds. This step is to enhance the hydrophilicity of the silicon wafer, allowing the electrolyte to more smoothly enter the bottom of the micro-pits, resulting in better electrodeposition effect.

[0091] Step S2.4, electrochemically depositing gold nanospike structures in a two-electrode system;

[0092] The step S2.4 is specifically as follows:

[0093] Step S2.4.1: Use a silicon wafer as the cathode (working electrode) and a carbon rod as the anode (auxiliary electrode). Use a copper rod-platinum electrode clamp to secure and connect to an electrochemical workstation. Preheat the electrochemical workstation for 30 minutes before electrodeposition.

[0094] In step S2.4.2, connect the green clip WE to the working electrode and the red clip CE to the auxiliary electrode. Select the i-tAmperometric iT Curve chronoamperometry method, set the potential to -3 V, the deposition time to 1800 s, the Sample Interval to 0.01 s, the Quiet Time to 0 s, and the Sensitivity to 1.e-003. Click OK to start electrodeposition.

[0095] Step S2.4.3: After the electrodeposition is completed, the sample is removed from the electrolytic cell and placed in an oven for drying;

[0096] The preparation of the gold-silicon SERS substrate with a micro-pit array on the surface is completed;

[0097] Figure 3 SEM image of gold-silicon SERS substrate;

[0098] The gold-silicon SERS substrate based on femtosecond laser processing of the present invention can be used to prepare a microfluidic chip, and the prepared microfluidic chip can be used to capture or detect cancer cells;

[0099] The cancer cells are PC-3 tumor cells and LNCaP tumor cells in prostate cancer;

[0100] in,

[0101] The application of gold-silicon SERS substrate processed by femtosecond laser in the detection of cancer cells includes the following steps:

[0102] The step S3 is specifically as follows:

[0103] The SERS substrate was treated with oxygen plasma for 90 to 120 seconds, and then soaked in a 10-6 mol / L MGITC solution in the dark for 3 to 4 hours. Finally, it was taken out and rinsed with deionized water to remove the free MGITC Raman reporter molecules not attached to the SERS substrate, and placed in a culture dish to air dry naturally. The prepared SERS substrate was placed on the test sample stage of the Renishaw Raman spectrometer, and then the SERS substrate was Raman detected and the results of the statistical test were collected. The test parameters of the Renishaw Raman spectrometer are: 20× microscope objective, excitation light source: 633nm, laser power: 1.7mW~5.5mW, integration time 1s, integration number 1 time, laser intensity 5%, and data from 10 random points on the SERS substrate were collected and recorded, and the average value was generated to compare the Raman signal with that of the ordinary gold-plated substrate. The results show that the gold-silicon SERS substrate processed by femtosecond laser has better Raman enhancement effect. Figure 4 As shown, cancer cells can also be captured Figure 5As shown in the figure, this is something that a smooth gold-plated substrate cannot achieve. The uniformity of SERS signals is one of the key indicators of a SERS substrate. SERS signals are detected at 10 random points on the same SERS substrate. Generally speaking, if the RSD (relative standard deviation) is below 10%, it means that the uniformity is good. Figure 6 As shown, the prepared SERS substrate was used to characterize the characteristic peak of Raman reporter molecule MGITC at 1614 cm -1 The peak value was used to measure the uniformity of the substrate, with RSD ≈ 7.49%, indicating good uniformity;

[0104] Figure 4 Raman signal detection of the prepared SERS substrate;

[0105] Step S4, substrate aptamer incubation and capture of cancer cells;

[0106] The step S4 is specifically as follows:

[0107] Step S4.1, modifying the two specific aptamers on the SERS substrate; first, the prepared SERS substrate is hydrophilicized, then placed in a solution of wy-5a and PSMA aptamers, slowly shaken for 24 hours, and finally, the residual aptamers are rinsed with buffer to complete the substrate modification;

[0108] Step S4.2, add 30 μL of 2.5% glutaraldehyde solution to the substrate for fixation for 2-3 hours; glutaraldehyde is used to fix the cancer cells and maintain their morphology when they are captured;

[0109] Step S4.3, placing the SERS substrate in 30%, 50%, and 80% alcohol solutions for 2 min each;

[0110] Step S4.4: Place the SERS substrate on a sample stage with an inclination angle of 45 degrees and spray palladium for 120-150 seconds.

[0111] The cells captured on the SERS substrate can then be observed using SEM.

[0112] Figure 5 SEM image of cells captured by SERS substrate;

[0113] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A method for preparing a gold-silicon SERS substrate based on femtosecond laser processing, characterized in that: The specific steps include: Step S1, femtosecond laser processing of silicon wafer; Step S1.1: Select a prime-grade single-polish silicon wafer with a resistance of 1-10Ω·cm and a thickness of 500±10μm. Step S1.2, cleaning the silicon wafer; Step S1.3, design the laser path and write the G code; Step S1.4, using femtosecond laser processing to process a micro-pit array on the surface of the silicon wafer; Step S2, electrochemically depositing gold nano-spike structures; Step S2.1, cutting and cleaning the silicon wafer; Step S2.2, preparing an electrolyte for electrochemical deposition; Step S2.3, treating the cleaned silicon wafer to make it hydrophilic; Step S2.4, electrochemically depositing gold nanospike structures in a two-electrode system; The preparation of gold-silicon SERS substrate with micro-pit array on the surface is completed. Step S3, using a confocal Raman spectrometer to test the uniformity of the substrate SERS performance; In step S4, the substrate aptamer is incubated and captures cancer cells.

2. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, characterized in that: The step S1.2 is specifically as follows: The silicon wafer was ultrasonically cleaned in deionized water and anhydrous ethanol for 10 minutes respectively, and then dried with nitrogen gas after cleaning.

3. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, characterized in that: The step S1.3 is specifically as follows: Step S1.3.1, design a micropit array, where the spacing between each micropit is approximately 10 μm, and each micropit is approximately 20 μm long, 20 μm wide, and 15 μm deep; Step S1.3.2, write the G code according to step S1.3.

1.

4. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, wherein: The step S1.4 is specifically as follows: Step S1.4.1: Fix the silicon wafer on a three-dimensional high-precision mobile platform, adjust the laser optical path, and assemble the objective lens to 10X / 0.25; Step S1.4.2: Turn on the CCD camera and focus on the silicon wafer plane until the image is clear; Step S1.4.3: After the image is clear, move the Z axis slightly to find the laser focus. The laser focus is the point where the laser spot is the smallest. In step S1.4.4, the power of the processing laser is 0.150 W, the frequency is 5.0 kHz, and the energy is 30.0 μJ. The G code is run to control the movement of the three-dimensional high-precision mobile platform while the femtosecond laser processes the silicon wafer.

5. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, characterized in that: The step S2.1 is specifically as follows: After the femtosecond laser processing, the silicon wafers were cut and ultrasonically cleaned in deionized water, anhydrous ethanol, and acetone for 15 minutes respectively. After cleaning, they were blown dry with nitrogen.

6. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, characterized in that: The step S2.2 is specifically as follows: preparing a 10 mM HAuCl4 solution and a 0.5 M HCl solution, adding equal volumes of the two solutions to the electrolytic cell, and then adding 1% Triton X-100 of the electrodeposition system to complete the electrolyte preparation.

7. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, characterized in that: The step S2.4 is specifically as follows: Step S2.4.1: Use a silicon wafer as the cathode (working electrode) and a carbon rod as the anode (auxiliary electrode). Use a copper rod-platinum electrode clamp to secure and connect to an electrochemical workstation. Preheat the electrochemical workstation for 30 minutes before electrodeposition. In step S2.4.2, connect the green clamp to the working electrode and the red clamp to the auxiliary electrode. Select chronoamperometry, set the potential to -3 V, and the deposition time to 1800 s. Step S2.4.3: After the electrodeposition is completed, the sample is removed from the electrolytic cell and placed in an oven for drying.

8. The method for preparing a gold-silicon SERS substrate based on femtosecond laser processing according to claim 1, wherein: The specific steps include: Step S3, immersing the substrate in 10 -6 mol / L MGITC solution for 10 to 12 hours, take out and air-dry before testing.

9. Use of the gold-silicon SERS substrate based on femtosecond laser processing according to any one of claims 1 to 8 in the detection of cancer cells.

10. The use of the gold-silicon SERS substrate based on femtosecond laser processing in the detection of cancer cells according to claim 9, characterized in that: The step S4 is specifically as follows: Step S4.1, modifying two specific aptamers on a SERS substrate; First, the prepared SERS substrate was hydrophilicized. Then, the treated SERS substrate was placed in a solution of wy-5a and PSMA aptamers and slowly shaken for 24 hours. Finally, the residual aptamers were washed away with buffer to complete the modification of the substrate. Step S4.2, add 30 μL of 2.5% glutaraldehyde solution to the substrate for fixation for 2-3 hours; Step S4.3, placing the SERS substrate in 30%, 50%, and 80% alcohol solutions for 2 min each; Step S4.4: Place the SERS substrate on a sample stage with an inclination angle of 45 degrees and spray palladium for 120-150 seconds. The cells captured on the SERS substrate can then be observed using SEM.

11. The use of the gold-silicon SERS substrate based on femtosecond laser processing in the detection of cancer cells according to claim 10, characterized in that: The cancer cells are PC-3 tumor cells and LNCaP tumor cells in prostate cancer.

12. Use of the gold-silicon SERS substrate based on femtosecond laser processing according to any one of claims 1 to 10 in cancer cell detection.

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