Gold monatomic group SERS (Surface Enhanced Raman Scattering) biosensor as well as preparation method and application thereof in beta amyloid protein detection

Abstract

The invention belongs to the technical field of biological detection, and relates to a gold single atom based SERS biosensor, a preparation method thereof and application of beta amyloid protein detection. The gold monatomic-based SERS biosensor comprises a silicon wafer substrate, a hydrophobic layer located on the surface of the silicon wafer and a gold monatomic-based SERS material layer located on the surface of the hydrophobic layer, and a gold monatomic-based SERS material is formed by loading gold monatomic on a nitrogen-doped carbon nanocage. The SERS sensor can realize detection of beta amyloid protein, is low in detection limit, and has great potential in detection of Alzheimer's disease.

Description

Technical Field

[0001] The present invention belongs to the technical field of biological detection, and relates to a gold single atom-based SERS biosensor, a preparation method thereof, and an application in beta-amyloid protein detection. Background Art

[0002] Alzheimer's disease (AD) is an irreversible neurodegenerative disorder that imposes a significant burden on the care of elderly people worldwide. Currently, the diagnosis of AD relies primarily on neuropsychological testing and imaging techniques. However, these psychological tests and cognitive assessments are significantly influenced by the patient's education and cultural background, which can lead to subjective and inaccurate test results. Positron emission tomography-based imaging technologies also face significant obstacles in achieving widespread adoption due to high hardware costs, resulting in prohibitively expensive tests. Therefore, an alternative approach using liquid biomarkers has emerged as an important detection strategy with the potential for large-scale and accurate detection of AD. Because the formation of amyloid-β inhibits ion channels, impairs calcium homeostasis, and impairs neuronal energy metabolism, ultimately leading to neuronal cell death, the detection of amyloid-β in cerebrospinal fluid and plasma provides a basis for the accurate diagnosis of AD.

[0003] Non-invasive and highly sensitive surface-enhanced Raman spectroscopy (SERS) is a powerful tool for material analysis, revealing the fingerprints of biomarkers at the cellular and molecular levels. Traditional SERS substrates primarily consist of noble metals and semiconductors. However, noble metal substrates have difficulty ensuring signal uniformity due to the presence of "hotspot" effects, while semiconductor substrates have limited signal enhancement capabilities. Therefore, the development of composite SERS substrates with high sensitivity, stability, selectivity, and reliability has become an urgent issue. In recent years, noble metal single-atom materials have shown broad application prospects in the materials field due to their high atomic efficiency and unique surface properties. In particular, research on SERS substrates from bulk to nanoclusters has been common, however, reports on single-atom SERS effects down to the atomic level are rare. The efficient interfacial charge transfer reactions of noble metal single atoms may provide promising candidate materials for the development of highly sensitive and stable SERS substrates. Summary of the Invention

[0004] In order to solve the above problems existing in the prior art, the purpose of the present invention is to provide a gold single atom-based SERS biosensor and its preparation method and β-amyloid protein detection application, so as to overcome the shortcomings of the prior art.

[0005] One object of the present invention is achieved by the following technical solutions:

[0006] A method for preparing a gold single atom-based SERS material comprises the following steps:

[0007] S1. Add alkaline solution to FeCl3·6H2O solution, maintain at 60-80℃, stir for 5-30min, and heat the obtained gel at 90-100℃ for 50-120h to obtain Fe2O3; add Fe2O3 and dopamine to alkaline aqueous solution, stir for 1-5h, collect, wash and dry to obtain Fe2O3@PDA, and calcine at 400-600℃ for 1-5h to obtain Fe3O4@NDC; add Fe3O4@NDC to acid solution, stir at 60-80℃ for 0.5-3h to remove the template, and obtain nitrogen-doped carbon nanocage (NDC) powder;

[0008] S2. NDC powder and cetyltrimethylammonium bromide are dispersed in water to obtain an NDC suspension. Subsequently, HAuCl4·4H2O solution is dropped into the NDC suspension. The mass ratio of HAuCl4·4H2O to NDC powder is 0.28-0.32:1. The mixture is stirred at 50-70°C for 8-16 hours, collected, washed, and dried, and then annealed at 70-90°C for 7-10 hours to obtain a gold single atom-loaded nitrogen-doped carbon nanocage material (Au1 / NDC).

[0009] In step S1:

[0010] The FeCl3·6H2O solution is formed by dissolving FeCl3·6H2O in water, and the concentration is 0.1-10 mol / L; the alkaline solution is formed by dissolving an alkali in water, the alkali is NaOH and / or KOH, and the concentration of the alkaline solution is 1-10 mol / L.

[0011] Preferably, the addition of the alkaline solution is controlled at a rate of 5 to 15 mL / min by a constant speed syringe pump.

[0012] Preferably, the molar ratio of FeCl3·6H2O to the alkali is 1:1 to 5. The volume of the alkali solution is the same as that of the FeCl3·6H2O solution.

[0013] Preferably, the mass ratio of Fe2O3 to dopamine is 1.2 to 5:1.

[0014] The alkaline aqueous solution may be an alkaline buffer solution, and an example of an alkaline buffer solution is Tris buffer solution.

[0015] Preferably, the calcination is carried out in an inert atmosphere, which may be nitrogen, argon, or the like.

[0016] The acid solution is formed by dissolving an acid in water, and the acid can be one or more of HCl solution, H2SO4 solution, HNO3 solution, hydrofluoric acid solution, etc. The concentration of the acid solution is 1 to 8 mol / L.

[0017] In step S2:

[0018] Preferably, the mass ratio of NDC powder to cetyltrimethylammonium bromide is 1:80-90.

[0019] Preferably, the annealing process is performed in a mixed atmosphere of hydrogen and argon (H2 / Ar).

[0020] The second object of the present invention is achieved through the following technical solutions:

[0021] A gold single atom-based SERS material is prepared by the preparation method provided in the first purpose.

[0022] The third object of the present invention is achieved through the following technical solutions:

[0023] A gold single atom-based SERS biosensor comprises a silicon wafer substrate, a hydrophobic layer on the surface of the silicon wafer, and a gold single atom-based SERS material layer on the surface of the hydrophobic layer, wherein the gold single atom-based SERS material layer comprises the above-mentioned gold single atom-based SERS material.

[0024] Preferably, the hydrophobic layer is prepared by a method comprising the following steps:

[0025] Alkyltrichlorosilane is mixed with water, and the mixture is added into an alkane solvent, then evenly dropped onto a silicon wafer and dried to obtain a hydrophobic layer.

[0026] Preferably, the gold single atom-based SERS material layer is prepared by a method comprising the following steps:

[0027] The gold single atom-based SERS material is dispersed in anhydrous ethanol, then dropped onto a silicon wafer having a hydrophobic layer, and dried to obtain a gold single atom-based SERS material layer.

[0028] The fourth object of the present invention is achieved through the following technical solutions:

[0029] A method for preparing a gold single atom-based SERS biosensor comprises the following steps:

[0030] (1) Alkyl trichlorosilane is mixed with water, the mixture is added to an alkane solvent, and then evenly dropped onto a silicon wafer and dried to obtain a hydrophobic layer;

[0031] (2) The gold single atom-based SERS material is dispersed in anhydrous ethanol, and then dropped onto the silicon wafer treated in step S1, and dried to obtain a gold single atom-based SERS biosensor.

[0032] Preferably, the alkyltrichlorosilane includes one or more of hexadecyltrichlorosilane, dodecyltrichlorosilane, octadecyltrichlorosilane, etc., and the alkane solvent includes one or more of pentane, hexane, heptane, octane, etc.

[0033] Preferably, the volume ratio of alkyltrichlorosilane to water is 30-80:1, and the volume ratio of the mixed liquid to the alkane solvent is 1:10-30.

[0034] Preferably, the step of mixing alkyltrichlorosilane with water comprises: adding water to alkyltrichlorosilane, vortexing at a speed of 1000-5000 rpm for 3-15 seconds, then ultrasonicating for 3-15 seconds, and repeating the vortexing and ultrasonicating steps 1-4 times to obtain a mixed solution.

[0035] Preferably, the mass ratio of the gold single atom-based SERS material to the volume of anhydrous ethanol is 1 mg: 2-10 ml.

[0036] The fifth object of the present invention is achieved through the following technical solutions:

[0037] Application of the gold single atom-based SERS biosensor in the detection of beta-amyloid protein.

[0038] Preferably, the application comprises the following steps: dropping a solution containing β-amyloid protein onto the gold single atom-based SERS biosensor, placing the solution under a micro Raman spectrometer after drying, and collecting a Raman spectrum.

[0039] Compared with the prior art, the present invention has the following beneficial effects:

[0040] 1. The synthesis of the new Au1 / NDC material can develop an atomic-scale SERS substrate, providing a new material basis for high-sensitivity SERS detection.

[0041] 2. The new Au1 / NDC material has high SERS sensitivity, and the SERS spectrum based on the Au1 / NDC substrate has good stability and reproducibility, with the lowest relative standard deviation of the characteristic peak intensity being 5.80%.

[0042] 3. The gold atoms in the Au1 / NDC material have high charge transfer activity and can more effectively promote the charge transfer from the material to the molecule than the gold clusters. Combined with the advantageous flat adsorption configuration of the MB molecules on the NDC surface, it provides a more effective charge transfer path for SERS detection, thereby enhancing the Raman signal.

[0043] 4. The SERS sensor prepared based on Au1 / NDC material can realize the detection of β-amyloid protein and has great potential in the detection of Alzheimer's disease.

[0044] 5. A hydrophobic layer is formed on the Au1 / NDC material-based SERS sensor. The hydrophobic layer is located between the silicon substrate and the Au1 / NDC material layer, which is beneficial to improving the detection sensitivity of the SERS sensor and can reduce the detection limit of β-amyloid protein from 10 -4 mol / L decreased to 10 -7 mol / L. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 These are transmission electron micrographs of the products of each step in Example 1, where A is Fe2O3 nanocubes, B is Fe2O3@PDA core-shell nanocubes, C is Fe3O4@NDC core-shell nanocubes, and D is nitrogen-doped carbon nanocage NDC.

[0046] Figure 2 These are spherical aberration electron micrographs of the gold single atoms supported on NDC in Example 1 and the gold clusters supported on NDC in Comparative Example 1, where A represents the gold single atom and B represents the gold cluster.

[0047] Figure 3 The SERS performance study of Au1 / NDC on dye molecules in Example 1, where AC represents NDC, Au1 / NDC and Au clu Comparison of the SERS effects of Au1 / NDC on methylene blue (MB) molecules, crystal violet (CV) molecules and copper phthalocyanine (CuPC). DF is the SERS detection limit of Au1 / NDC for MB, CV and CuPC molecules. GI is the SERS detection stability of Au1 / NDC for MB, CV and CuPC molecules.

[0048] Figure 4 This is the crystal diffraction pattern of MB molecules adsorbed on the surface of carbon analogues.

[0049] Figure 5 This is a comparison diagram of the crystal configuration energy of MB molecules adsorbed on the NDC surface.

[0050] Figure 6 The Au1 / NDC of Example 1 and the Au of Comparative Example 1 clu Comparison of the number of charge transfers for / NDC.

[0051] Figure 7 This is a scanning electron microscope image of the hydrophobic surface of the hydrophobic layer prepared in Example 2.

[0052] Figure 8 This is a comparison chart of the detection limits of the SERS sensors of Example 2 and Comparative Example 2 for detecting amyloid β.

[0053] Figure 9 Fluorescence, bright field and Merge images of fluorescently modified β-amyloid protein.

[0054] Figure 10 These are the Raman bright field, selected point and average spectra of β-amyloid protein. DETAILED DESCRIPTION

[0055] Below by specific embodiment and accompanying drawing, technical scheme of the present invention is further described explanation, it should be understood that specific embodiment described herein is only for helping to understand the present invention, is not used for specific limitation of the present invention.And accompanying drawing used herein, is only for better illustrating the disclosure of the present invention, does not have limiting effect on protection scope.If no special instructions, the raw materials adopted in the embodiment of the present invention are all raw materials commonly used in this area, and the method adopted in the embodiment is all conventional method in this area.

[0056] Example 1

[0057] The gold single atom-based SERS material provided in this embodiment was prepared by the following method:

[0058] S1. Take 25mL of FeCl3·6H2O solution (2M) and stir it in a 75℃ oil bath for 10 minutes; then, put 25mL of NaOH solution (5.4M) into a plastic syringe and connect it to a constant speed injection pump to control the addition of NaOH at a rate of 10mL / min. After the addition is complete, maintain the temperature and stir for further 10 minutes. During this period, the reactant undergoes a phase transition from liquid to gel. The obtained gel is placed in an oven and reacted at 100℃ for 96h. The red precipitate is collected and washed with water and ethanol several times. After drying, Fe2O3 nanocubic powder (such as Figure 1 A). Subsequently, 160 mg of Fe2O3 was dispersed in 200 mL of 10 mM Tris buffer solution, and the resulting suspension was stirred with 80 mg of dopamine for 3 h. The Fe2O3@PDA core-shell nanocube powder (e.g. Figure 1 B). The collected Fe2O3@PDA was calcined in a tube furnace at 500℃ under argon atmosphere for 3h to obtain a black Fe3O4@NDC composite material (such as Figure 1 C). Next, the Fe3O4@NDC composite material was dispersed in a strong acidic solution (HCl, 4M, 100 mL) and mechanically stirred (70°C, 250 rpm) for 1 h to remove the template. The product was washed and dried to obtain nitrogen-doped carbon nanocage (NDC) powder (such as Figure 1 D).

[0059] S2. Disperse 3 mg of NDC powder obtained in step S1 and 260 mg of cetyltrimethylammonium bromide in 80 mL of deionized water. Ultrasonicate for 5 minutes and stir for 30 minutes to evenly disperse the NDC particles to obtain an NDC suspension. Subsequently, 0.9 mL of 1.0 mg / mL HAuCl4·4H2O was dropwise added to the NDC suspension under continuous stirring at 60°C. After stirring for 10 hours, the sample was collected and washed several times. The dried powder was annealed at 80°C in a 5% hydrogen and argon (H2 / Ar) mixed atmosphere for 8 hours to obtain the gold single-atom-loaded nitrogen-doped carbon nanocage material (Au1 / NDC).

[0060] After sonication, the distribution of gold atoms was observed under a spherical aberration electron microscope ( Figure 2 A). Due to the unique structure of NDC, gold single atoms loaded on NDC have better dispersion, which is beneficial to improving the SERS intensity.

[0061] Comparative Example 1

[0062] The difference between Comparative Example 1 and Example 1 is that in step S2 of Comparative Example 1, the amount of HAuCl4·4H2O added is increased to 1.1 mL. The other steps are the same as those in Example 1. The obtained product is nitrogen-doped carbon nanocages Au loaded with gold clusters. clu / NDC, the distribution of gold clusters was observed under spherical aberration electron microscope after ultrasonic disruption ( Figure 2 B).

[0063] 1. Study on the SERS performance of Au1 / NDC on dye molecules

[0064] In order to illustrate the improvement of the SERS performance of NDC carbon-based materials by gold single atoms and the high atomic efficiency of gold single atoms, the NDC prepared in step S1 of Example 1, the Au1 / NDC of Example 1 and the Au clu / NDC were used to compare the SERS spectra of dye molecules. -4 mol / L, Au1 / NDC can enhance the signal up to 5.5 times compared with NDC, while the formation of gold clusters weakens the signal enhancement ability ( Figure 3 A), similarly, when the concentration of crystal violet (CV) molecules is 10 -4 mol / L, Au1 / NDC can enhance the signal up to 6 times compared with NDC, while the formation of gold clusters weakens the signal enhancement ability ( Figure 3 B); when the concentration of copper phthalocyanine (CuPC) molecules is 10 -5 mol / L, Au1 / NDC can enhance the signal up to 4.25 times compared with NDC, while the formation of gold clusters weakens the signal enhancement ability ( Figure 3 C).

[0065] In order to illustrate the sensitivity of Au1 / NDC, the detection limits of MB, CV, and CuPC molecules were analyzed. It can be seen that the lowest concentration that Au1 / NDC can detect is 10 -7 mol / L of MB( Figure 3 D); the lowest detectable concentration of Au1 / NDC is 10 -8 mol / L of MB( Figure 3 E); the lowest detectable concentration of Au1 / NDC is 10 -6 mol / L CuPC( Figure 3 F).

[0066] In order to demonstrate the good stability and reproducibility of the SERS spectra based on the Au1 / NDC substrate, multiple Raman spectra of Au1 / NDC detecting MB (Au1 / NDC-MB), Au1 / NDC detecting CV (Au1 / NDC-CV), and Au1 / NDC detecting CuPC (Au1 / NDC-CuPC) were collected. Figure 3 The GI is 20 SERS spectra, and the relative standard deviation (RSD) of the characteristic peak intensity is calculated, with the lowest being 5.80%. The above results show that the SERS spectrum based on Au1 / NDC substrate has better stability and reproducibility.

[0067] 2. Study on the dominant adsorption configuration of the molecule to be tested on NDC

[0068] In order to explore the dominant adsorption configuration of the test molecules on NDC, a systematic study was conducted combining experimental characterization and theoretical calculations. First, the orientation distribution of MB molecules on the carbon material surface was analyzed using grazing incidence wide-angle X-ray scattering. The results showed that MB molecules exhibited a highly ordered planar configuration ( Figure 4 Further density functional theory calculations were used to compare the two adsorption configurations of MB molecules on NDC (lying vs. standing). It was found that the absolute value of the adsorption energy of the lying configuration (-14585.90 eV) was higher than that of the standing configuration (-14585.58 eV), indicating that MB molecules tend to be adsorbed on the NDC surface in a planar manner ( Figure 5 This flat configuration can provide a more efficient charge transfer path for SERS detection, thereby enhancing the Raman signal.

[0069] 3. Study on the enhancement mechanism of Au1 / NDC substrate

[0070] In order to clarify the enhancement mechanism of the Au1 / NDC substrate, MB was constructed on Au1 / NDC and Au clu / The density functional theory model on NDC was constructed, that is, Au1 / NDC-MB and Au clu / NDC-MB. By analyzing the difference in electron density of MB adsorbed on different SERS materials, it was found that the rate of charge transfer promoted by a single Au atom from the material to the molecule was 0.236e( Figure 6 ), which indicates that Au1 / NDC has a high charge transfer activity, while the promotion effect of Au clusters on charge transfer is only 0.054e, indicating that its charge transfer ability is weaker than that of a single Au atom, which is consistent with the above experimental results.

[0071] Example 2

[0072] The gold single atom-based SERS biosensor provided in Example 2 was prepared by the following method:

[0073] (1) Take 1 mL of octadecyltrichlorosilane, add 20 μL of deionized water, vortex at 3000 rpm, and sonicate for 10 s. Repeat the vortexing and sonication steps once. Then, take 500 μL and add 10 mL of n-hexane. Shake evenly and drop it onto a clean silicon wafer. Dry it to form a hydrophobic layer.

[0074] (2) 2 mg of Au1 / NDC prepared in Example 1 was dispersed in 10 ml of anhydrous ethanol, and 20 μL was dropped onto the clean silicon wafer treated in step (1), and allowed to dry naturally to obtain a gold single atom-based SERS biosensor.

[0075] The surface scanning electron microscope of the hydrophobic layer formed in Example 2 is as follows Figure 7 As shown, its microstructure surface texture is periodically arranged, and these textures are composed of a combination of protrusions and grooves with sizes ranging from submicron to nanometer levels.

[0076] Comparative Example 2

[0077] The gold single atom-based SERS biosensor provided in Comparative Example 2 was prepared by the following method: 2 mg of Au1 / NDC prepared in Example 1 was dispersed in 10 ml of anhydrous ethanol, 20 μL was dropped on a clean silicon wafer, and allowed to stand and dry naturally to obtain a gold single atom-based SERS biosensor.

[0078] 20 μL of the prepared β-amyloid protein solution was dropped onto the biosensors of Example 2 and Comparative Example 2, respectively, and allowed to dry naturally. After drying, the biosensors were placed under a micro-Raman spectrometer for Raman detection. The SERS sensor without a hydrophobic layer can detect 10 -4 mol / L of β-amyloid protein, and the SERS sensor with a hydrophobic layer can detect 10 -7 mol / L of β-amyloid protein ( Figure 8 ). By forming a hydrophobic layer on the SERS sensor, it is beneficial to improve the detection sensitivity of the SERS sensor and reduce the detection limit of β-amyloid protein.

[0079] In order to demonstrate that the collected Raman spectra of amyloid-β were not interfered by the probe substrate or the environment, a fluorescence-Raman co-localization experiment was performed on amyloid-β labeled with fluorescein isothiocyanate (FITC). First, the amyloid-β was fixed on a glass plate with region markings, and its fluorescence signal was collected under a fluorescence microscope to determine the fluorescence signal region ( Figure 9 ); Single-point Raman signal acquisition of amyloid beta was performed in the calibration area, and the acquired spectrum was compared with the spectrum of Example 6 to confirm that the pattern was a characteristic spectrum of amyloid beta ( Figure 10 ).

[0080] The various aspects, embodiments, and features of the present invention should be considered in all respects as illustrative and not limiting, the scope of which is defined solely by the claims. Other embodiments, modifications, and uses will be apparent to those skilled in the art without departing from the spirit and scope of the invention as claimed.

[0081] In the preparation method of the present invention, the order of the steps is not limited to the order listed. Persons skilled in the art will appreciate that variations in the order of the steps are within the scope of the present invention without inventive effort. Furthermore, two or more steps or actions may be performed simultaneously.

[0082] Finally, it should be noted that the specific embodiments described herein are merely illustrative of the present invention and are not intended to limit its implementation. Persons skilled in the art may make various modifications, additions, or substitute similar methods for the described specific embodiments. It is not necessary and impossible to provide comprehensive examples of all implementations here. However, obvious variations or modifications arising from the essential spirit of the present invention remain within the scope of protection of the present invention, and interpreting them as any additional limitations would be contrary to the spirit of the present invention.

Claims

1. A method for preparing a gold single atom-based SERS material, characterized in that: The following steps are involved: S1. Add alkaline solution to FeCl3·6H2O solution, maintain at 60-80℃, stir for 5-30min, and heat the obtained gel at 90-100℃ for 50-120h to obtain Fe2O3; add Fe2O3 and dopamine to alkaline aqueous solution, stir for 1-5h, collect, wash and dry to obtain Fe2O3@PDA, and calcine at 400-600℃ for 1-5h to obtain Fe3O4@NDC; add Fe3O4@NDC to acid solution, stir at 60-80℃ for 0.5-3h to remove the template, and obtain nitrogen-doped carbon nanocage powder; S2. NDC powder and cetyltrimethylammonium bromide are dispersed in water to obtain an NDC suspension. Subsequently, HAuCl4·4H2O solution is dropped into the NDC suspension. The mass ratio of HAuCl4·4H2O to NDC powder is 0.28-0.32:

1. The mixture is stirred at 50-70°C for 8-16 hours, collected, washed, and dried, and then annealed at 70-90°C for 7-10 hours to obtain a gold single atom-loaded nitrogen-doped carbon nanocage material.

2. The preparation method according to claim 1, characterized in that In step S1, the FeCl3·6H2O solution is formed by dissolving FeCl3·6H2O in water, and the concentration is 0.1-10 mol / L; the alkaline solution is formed by dissolving an alkali in water, the alkali is NaOH and / or KOH, and the concentration of the alkaline solution is 1-10 mol / L; and / or, controlling the addition of the alkaline solution at a rate of 5 to 15 mL / min using a constant-speed syringe pump; and / or, the molar ratio of FeCl3·6H2O to the alkali is 1:1 to 5; and / or, the mass ratio of Fe2O3 to dopamine is 1.2 to 5:1; and / or, calcination is carried out in an inert atmosphere; And / or, the acid solution is formed by dissolving an acid in water, the acid is one or more of HCl solution, H2SO4 solution, HNO3 solution, and hydrofluoric acid solution, and the concentration of the acid solution is 1 to 8 mol / L.

3. The preparation method according to claim 1, characterized in that In step S2, the mass ratio of NDC powder to cetyltrimethylammonium bromide is 1:80-90; And / or, the annealing process is performed in a mixed atmosphere of hydrogen and argon.

4. A gold single atom-based SERS material, characterized in that: The gold single atom-based SERS material is prepared by the preparation method described in any one of claims 1 to 3.

5. A gold single atom-based SERS biosensor, characterized in that: The invention comprises a silicon wafer substrate, a hydrophobic layer located on the surface of the silicon wafer, and a gold single atom-based SERS material layer located on the surface of the hydrophobic layer, wherein the gold single atom-based SERS material layer comprises the gold single atom-based SERS material as claimed in claim 4 or the gold single atom-based SERS material prepared by any one of the preparation methods as claimed in claims 1 to 3.

6. The gold single atom-based SERS biosensor according to claim 5, characterized in that: The hydrophobic layer is prepared by a method comprising the following steps: Alkyl trichlorosilane is mixed with water, the mixture is added to an alkane solvent, and then evenly dropped onto a silicon wafer and dried to obtain a hydrophobic layer; And / or, the gold single atom-based SERS material layer is prepared by a method comprising the following steps: The gold single atom-based SERS material is dispersed in anhydrous ethanol, then dropped onto a silicon wafer having a hydrophobic layer, and dried to obtain a gold single atom-based SERS material layer.

7. A method for preparing a gold single atom-based SERS biosensor, characterized in that: The following steps are involved: (1) Alkyl trichlorosilane is mixed with water, the mixture is added to an alkane solvent, and then evenly dropped onto a silicon wafer and dried to obtain a hydrophobic layer; (2) The gold single atom-based SERS material is dispersed in anhydrous ethanol, and then dropped onto the silicon wafer treated in step S1, and dried to obtain a gold single atom-based SERS biosensor.

8. The preparation method according to claim 7, characterized in that The alkyltrichlorosilane includes one or more of hexadecyltrichlorosilane, dodecyltrichlorosilane, and octadecyltrichlorosilane; and / or, the alkane solvent includes one or more of pentane, hexane, heptane, octane, etc.; and / or, the volume ratio of alkyltrichlorosilane to water is 30 to 80:1, and the volume ratio of the mixed solution to the alkane solvent is 1:10 to 30; And / or, the mass ratio of the gold single atom-based SERS material to the volume of anhydrous ethanol is 1 mg: 2-10 ml.

9. The preparation method according to claim 7, characterized in that The step of mixing alkyltrichlorosilane with water comprises: adding water into alkyltrichlorosilane, vortexing at a speed of 1000-5000 rpm for 3-15 seconds, then ultrasonically treating for 3-15 seconds, and repeating the vortexing and ultrasonicating steps 1-4 times to obtain a mixed solution.

10. Use of the gold single atom-based SERS biosensor according to claim 5 in the detection of β-amyloid protein.

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