Surface-enhanced Raman substrate based on gold nanorod self-assembly and nanosphere photoetching as well as preparation method and application of surface-enhanced Raman substrate

Abstract

The invention discloses a surface-enhanced Raman substrate based on gold nanorod self-assembly and nanosphere photoetching as well as a preparation method and application of the surface-enhanced Raman substrate, and belongs to the technical field of surface-enhanced Raman substrates. A large-area single-layer nanosphere template is prepared from 500nm silicon dioxide microspheres through a Langmuir-Blodgett method, the SERS substrate with high-density ordered hot spots is constructed by optimizing nanosphere photoetching parameters and self-assembly parameters of gold nanorods, the performance limitation of a traditional substrate is broken through, and a new thought is provided for development of a high-performance SERS device.

Description

Technical Field

[0001] The present invention belongs to the technical field of surface-enhanced Raman substrates, and particularly relates to a surface-enhanced Raman substrate based on self-assembly of gold nanorods and nanosphere lithography, and a preparation method and application thereof. Background Art

[0002] Surface-Enhanced Raman Spectroscopy (SERS), as a highly sensitive molecular detection technology, has shown important application values in the fields of trace substance detection, biomedical diagnosis, environmental monitoring, food safety, etc. The core of the SERS technology relies on nanostructured substrates with local surface plasmon resonance (LSPR) effect, and the "hot spots" formed on their surfaces are the key to signal enhancement. However, the existing SERS substrates still face significant challenges in terms of sensitivity, stability, reproducibility and preparation cost, and it is difficult to achieve large-area preparation, which restricts their practical applications. Traditional SERS substrates mostly adopt randomly distributed noble metal nanoparticles (such as gold and silver nanospheres), and are obtained by chemical reduction method or physical sputtering method. Although such substrates are simple to prepare, the random aggregation of nanoparticles results in uneven distribution of "hot spots" and poor repeatability between batches, making it difficult to meet the requirements of quantitative analysis. To improve the performance of the substrates, researchers have turned to the controllable construction of ordered nanostructures. For example, electron beam lithography (EBL) and focused ion beam etching (FIB) technologies can precisely design periodic nanoarrays, but their equipment costs are high, the processes are complex, and it is difficult to achieve large-area preparation.

[0003] In recent years, Nanosphere Lithography (NSL) technology has attracted attention due to its advantages of low cost, high throughput and scalability. This method uses self-assembled monolayer nanospheres as a mask, and combines metal deposition or etching processes to prepare periodic nanostructures. However, the structures formed by NSL have high symmetry and low geometric degrees of freedom, resulting in limited hot spot density and electromagnetic field enhancement factor. In addition, the monolayer nanosphere template is easily affected by factors such as the hydrophilicity / hydrophobicity of the substrate, environmental temperature and humidity, and it is difficult to achieve large-area uniform assembly, further restricting the performance uniformity of the substrate.

[0004] On the other hand, anisotropic metal nanomaterials (such as gold nanorods) have become a research hotspot for SERS substrates due to their unique LSPR characteristics. The aspect ratio of gold nanorods can adjust their longitudinal and transverse plasma resonance wavelengths, thereby achieving strong electromagnetic field enhancement in the near-infrared region and broadening the application spectral range of SERS. However, the self-assembly process of gold nanorods is limited by their dispersion stability and orientation randomness, making it difficult to form a high-density hotspot with regular arrangement. In the existing technology, although external field induction (such as electric field, magnetic field) or template-assisted assembly can improve the arrangement order, the process is often complex or requires the introduction of additional functionalization modifications, resulting in increased costs and possible introduction of impurity interference. Summary of the Invention

[0005] Aiming at the problems of insufficient sensitivity, uneven hotspot distribution, high preparation cost and poor reproducibility of the existing surface-enhanced Raman spectroscopy (SERS) substrates, the present invention provides a SERS substrate based on the self-assembly of gold nanorods and nanosphere lithography, as well as its preparation method and application. By combining the periodic template advantages of nanosphere lithography with the directional arrangement characteristics of gold nanorods, a high-density ordered electromagnetic "hotspot" array is constructed, realizing the high sensitivity, high reproducibility, low cost and scalable preparation of the SERS substrate, and enhancing its application potential in the fields of trace detection, biosensing, etc.

[0006] To achieve the above object, the present invention provides the following technical solutions:

[0007] One of the technical solutions of the present invention:

[0008] The present invention provides a preparation method of a surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography, including the following steps:

[0009] Prepare a monolayer SiO2 microsphere film by the Langmuir-Blodgett method, transfer the monolayer SiO2 microsphere film to the surface of a silicon wafer, and form a moth-eye biomimetic nanoarray by reactive ion etching. The reactive ions are a mixture of SF6, He and O2;

[0010] Prepare a suspension containing gold nanorods by the seed growth method. During the preparation of the suspension containing gold nanorods, cetyltrimethylammonium bromide (CTAB) and sodium oleate are added, and a gold nanorod film is formed by liquid-liquid interface self-assembly;

[0011] Transfer the gold nanorod film to the surface of the moth-eye biomimetic nanoarray to obtain the surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography.

[0012] Further, the diameter of the SiO2 microspheres is 500 nm.

[0013] The present invention combines the anisotropic LSPR characteristics of gold nanorods with nanosphere lithography technology. A large-area monolayer nanosphere template is prepared by using 500-nm silica microspheres through the Langmuir-Blodgett method. By optimizing the nanosphere lithography parameters and the self-assembly parameters of gold nanorods, a SERS substrate with high-density ordered hot spots is constructed, breaking through the performance limitations of traditional substrates and providing new ideas for the development of high-performance SERS devices.

[0014] Further, the steps for preparing a monolayer SiO2 film by the Langmuir-Blodgett method are as follows: centrifuging the SiO2 microsphere suspension, discarding the supernatant, dispersing the precipitate in ethanol, and ultrasonically treating until completely dispersed to obtain a SiO2 microsphere suspension free of impurities; using the SiO2 microsphere suspension free of impurities to assemble a monolayer SiO2 microsphere film on the surface of a glass slide.

[0015] Further, the steps for transferring the monolayer SiO2 microsphere film to the surface of a silicon wafer are as follows: immersing the silicon wafer in a piranha solution to obtain a pretreated silicon wafer; transferring the monolayer SiO2 microsphere film to the surface of the pretreated silicon wafer and drying.

[0016] Further, the steps for forming a moth-eye biomimetic nanoarray by reactive ion etching are as follows: placing the silicon wafer covered with the monolayer SiO2 microsphere film in a reactive ion etching device, introducing a mixed gas of SF6, He, and O2, with the radio frequency power being 50% (indicating that the etching power is 50% of the maximum rated power of the radio frequency power supply of the device), and the etching time being 200 - 300 s, preferably 200 s.

[0017] Further, in the step of forming a moth-eye biomimetic nanoarray by reactive ion etching, a mixed gas of SF6 35 sccm, He 35 sccm, and O2 5 sccm is introduced.

[0018] Further, the steps for preparing the gold nanorod film are as follows:

[0019] a. Mixing a HAuCl4 solution with a cetyltrimethylammonium bromide solution, adding a NaBH4 solution, stirring evenly, and aging at room temperature to obtain a gold nanorod seed solution;

[0020] b. Mixing cetyltrimethylammonium bromide and sodium oleate in water, adding an AgNO3 solution, standing, adding a HAuCl4 solution, stirring until colorless, adjusting the pH to 1 - 2, adding ascorbic acid, stirring evenly, adding the gold nanorod seed solution, and standing to obtain a gold nanorod suspension;

[0021] c. Centrifuge the gold nanorod suspension, incubate it first with a sodium polystyrene sulfonate (Na-PSS) solution, then centrifuge and wash it with a trisodium citrate solution, and concentrate it to obtain a concentrated solution;

[0022] d. Mix the concentrated solution with dichloromethane in a centrifuge tube, shake it and then let it stand, and form an orderly arranged gold nanorod film at the water-air interface.

[0023] Further, in step b, the mass ratio of cetyltrimethylammonium bromide to sodium oleate is (5-7):1, preferably 5.7:1.

[0024] Further, in step c, the concentration of the Na-PSS solution is 0.05-0.4 wt%. Exemplarily, the concentration of the Na-PSS solution is 0.05 wt%, 0.1 wt%, 0.2 wt% or 0.4 wt%, preferably 0.2 wt%.

[0025] Further, in step c, the concentration of the trisodium citrate solution is 2-20 mM. Exemplarily, the concentration of the trisodium citrate solution is 2 mM, 4 mM, 10 mM or 20 mM, preferably 10 mM.

[0026] The present invention is based on the liquid-liquid interface self-assembly technology. Through the interfacial tension gradient, gold nanorods (GNRs) form a uniform, dense and renewable self-assembled monolayer film at the air-water interface, and are transferred to the surface of the nanoarray structure formed by etching to construct a high-performance SERS substrate. By mixing dichloromethane solution (CH2Cl2 solution) with the gold nanorod suspension, due to the different densities between CH2Cl2 and the GNRs suspension, the GNRs are driven by the interfacial tension gradient to aggregate at the air-water interface, forming a neatly arranged monolayer gold nanorod film (gold film). When the gold film is transferred to the silicon wafer, due to the hydrophilicity of the silicon wafer, the gold film can be adsorbed on the silicon wafer surface autonomously; the biomimetic moth-eye array structure formed on the silicon wafer surface by ion etching further provides a capillary force gradient to attract the gold nanorods into the structure, promoting the close packing of GNRs, forming a large number of plasmonic hot spots to enhance the Raman signal. At the same time, under the action of the electrostatic force and van der Waals force between the biomimetic moth-eye array structure and the gold film, a uniformly arranged gold nanorod film is formed on the structure.

[0027] The second technical solution of the present invention:

[0028] The present invention also provides a surface-enhanced Raman substrate based on gold nanorod self-assembly and nanosphere lithography prepared according to the above method, which has a biomimetic moth-eye nanoarray, and gold nanorods are present between the biomimetic moth-eye nanoarrays.

[0029] The present invention provides a surface-enhanced Raman substrate (SERS substrate) based on the self-assembly of gold nanorods and nanosphere lithography technology. The SERS substrate has an array structure formed by an array of gold nanorods and a reactive ion-etched silicon wafer. Compared with other SERS substrates, the SERS substrate prepared by the method of the present invention has an existence of a moth-eye biomimetic array structure, enabling the gold nanorods to exist in the structural gaps, providing more adsorption sites for the gold nanorods, and greatly increasing the adsorption stability. At the same time, the SERS performance can be improved by increasing the number of assembly layers of the gold nanorod thin film on the structure.

[0030] The third technical solution of the present invention:

[0031] The present invention also provides an application of the surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography in the field of Raman sensing.

[0032] Compared with the prior art, the present invention has the following advantages and technical effects:

[0033] The surface-enhanced Raman spectrum and its preparation method based on the self-assembly of gold nanorods and nanosphere lithography provided by the present invention introduce gold nanorods into the gaps of the periodic array structure of the silicon substrate through the combination of nanosphere lithography and the self-assembly of gold nanorods, forming multiple electromagnetic field enhancement regions, obtaining uniform "hot spots" while providing a stable solid support to increase the stability of the substrate.

[0034] The preparation method of the present invention is simple in operation, low in preparation cost, can realize large-area batch production of the substrate, and the obtained substrate has strong stability, good repeatability, and high sensitivity. Description of the Drawings

[0035] The drawings forming a part of the present invention are used to provide a further understanding of the present invention. The schematic embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute an improper limitation to the present invention. In the drawings:

[0036] Figure 1 It is a self-assembly device diagram of the SiO2 microsphere thin film in Example 1;

[0037] Figure 2 It is a scanning electron microscope image of the monolayer SiO2 microsphere mask formed by self-assembly in Example 1;

[0038] Figure 3 It is a scanning electron microscope image of the nanoscale arrays formed with different etching times in Examples 1-2 and Comparative Examples 1-2. a is Example 1 (etching time is 200 s), b is Example 2 (etching time is 300 s), c is Comparative Example 1 (etching time is 400 s), and d is Comparative Example 2 (etching time is 500 s);

[0039] Figure 4 SEM image of the gold nanorods prepared in Example 3;

[0040] Figure 5 SEM images of gold nanorod arrays prepared with different concentration multiples of gold nanosuspensions, where the concentration of Na-PSS is 0.05% (mass concentration) and the concentration of trisodium citrate solution is 4 mM. Among them, a and b are 2 times (Example 4), c and d are 10 times (Example 3), and e and f are 20 times (Example 5);

[0041] Figure 6 SEM images of gold nanorod arrays prepared with different concentrations of trisodium citrate solutions, where the concentration multiple of the gold nanorod suspension is 10 times and the concentration of Na-PSS is 0.05% (mass concentration). a and b are 2 mM (Example 6), c and d are 10 mM (Example 7), and e and f are 20 mM (Example 8);

[0042] Figure 7 SEM images of gold nanorod arrays prepared with different concentrations of Na-PSS, where the concentration multiple of the gold nanorod suspension is 10 times and the concentration of trisodium citrate solution is 10 mM. a and b are 0.1% (Example 9), c and d are 0.2% (Example 10), and e and f are 0.4% (Example 11), all in mass concentration;

[0043] Figure 8 SEM images of surface-enhanced Raman scattering substrates (abbreviated as SERS substrates) based on self-assembled gold nanorods and nanosphere lithography, prepared with different numbers of self-assembled gold nanorod thin films on silicon wafers and moth-eye bionic array structures in Examples 12 to 14. a and d are 1 layer, b and e are 2 layers, and c and f are 4 layers; a, b, c are moth-eye bionic array structures, and b, e, f are silicon wafers.

[0044] Figure 9 Raman spectra of substrates with different self-assembled layers on silicon wafers and moth-eye bionic array structures in Examples 12 to 14;

[0045] Figure 10 Results of the stability differences between the substrate (SERS substrate prepared in Example 12) and the silicon wafer (control in Example 12) after the soaking experiment and the dropping experiment. a is after dropping pure water on the SERS substrate; b is after dropping pure water on a single self-assembled structure; c is after soaking the SERS substrate in pure water; d is after soaking a single self-assembled structure in pure water;

[0046] Figure 11 Curves showing the variation of the Raman signal intensity of the signal molecule R6G with concentration on the substrates in Examples 12 to 14 (10 -12 ~10 -6 M);

[0047] Figure 12 For the linear fitting of the Raman signal intensity of R6G signal molecules on the substrate in Example 12 with the change of concentration;

[0048] Figure 13 For the repeatability analysis of the Raman spectra at 20 random detection points on the substrate in Example 12;

[0049] Figure 14 For the large - area distribution of self - assembled gold nanorods on the silicon wafer in Example 10. Detailed implementation manners

[0050] Now, various exemplary implementation manners of the present invention will be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and implementation schemes of the present invention.

[0051] It should be understood that the terms described in the present invention are only for describing specific implementation manners and are not used to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Each intermediate value within any stated value or stated range, as well as each smaller range between any other stated value or intermediate value within the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range.

[0052] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention pertains. Although the present invention only describes preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In case of conflict with any incorporated document, the content of this specification shall prevail.

[0053] Without departing from the scope or spirit of the present invention, various improvements and changes can be made to the specific implementation manners of the present invention specification, which are obvious to those skilled in the art. Other implementation manners obtained from the specification of the present invention are obvious to those skilled in the art. The specification and examples of the present invention are only exemplary.

[0054] Regarding the terms "comprising", "including", "having", "containing", etc. used herein, they are all open - ended terms, meaning including but not limited to.

[0055] The present invention obtains a monodisperse SiO2 microsphere suspension through centrifugal cleaning; forms a monolayer film by the Langmuir-Blodgett method and transfers it to a silicon wafer; and forms a nanoscale array by reactive ion etching (a mixed gas of SF6 / He / O2). By using the above method, a large-area self-assembled monolayer mask of SiO2 can be prepared, solving the problems of difficult large-area preparation and difficult homogeneity realization in nanosphere lithography technology.

[0056] The present invention prepares gold nanorods with a high aspect ratio by a binary surfactant seed growth method using cetyltrimethylammonium bromide (CTAB) and sodium oleate (NaOL); forms a dense monolayer film through liquid-liquid interface self-assembly; and enhances stability through surface ligand replacement (CTAB → sodium polystyrenesulfonate (Na-PSS) → trisodium citrate). The interfacial tension is used to drive the gold nanorods to form a dense monolayer film, and the close and orderly self-assembly of the monolayer gold nanorods is achieved by adjusting the self-assembly parameters. Compared with other methods, large-area preparation is realized while avoiding aggregation.

[0057] The present invention transfers the gold nanorod film to the surface of an artificial moth-eye array structure. Due to the capillary force generated by the artificial moth-eye array structure on the substrate surface, the gold nanorods enter the structure, further increasing the density of the gold nanorods. At the same time, due to the electrostatic interaction and van der Waals force between the array structure and the gold nanorods, the stability of the gold nanorods on the substrate is increased, so that the gold nanorods are not easily piled up while being affected by the liquid surface tension, and still have good uniformity.

[0058] In the following examples and comparative examples of the present invention, the reactive ion etching equipment is an RIE etching machine (Filmlab-R100), and the maximum rated power of the radio frequency power supply is 150W.

[0059] Unless otherwise specified, the room temperature in the present invention is uniformly calculated as 25 ± 2°C.

[0060] All raw materials used in the examples of the present invention are obtained by purchasing commercially. As an example, the SiO2 microsphere suspension with a particle size of 500 nm is purchased from Macklin Inc (Macklin Reagent).

[0061] It should be noted that the parts not described in detail in the present invention are all conventional operation means in the art and are not the focus of the present invention.

[0062] The technical solution of the present invention is further described below through examples.

[0063] Example 1

[0064] This example provides a method for preparing an artificial moth-eye nanoscale array:

[0065] Step 1: Cleaning of SiO2 microspheres

[0066] Take 200 μL of the SiO2 microsphere suspension with a particle size of 500 nm, centrifuge it at 16,000 rpm for 15 min, discard the supernatant, disperse the precipitate in 300 μL of ethanol, and ultrasonicate for 30 min until completely dispersed. Repeat the washing 3 times to remove impurities.

[0067] Step 2: Pretreatment of the silicon wafer

[0068] Immerse a 1 cm × 1 cm silicon wafer into the piranha solution (concentrated sulfuric acid (mass content is 95%, the same below): hydrogen peroxide = 2:1, volume ratio), and treat it at 120 °C for 50 min to remove surface organic matters and oxides.

[0069] Step 3: Large-area assembly of monolayer SiO2 microspheres

[0070] Inject the SiO2 microsphere suspension after removing impurities into a petri dish containing 2 / 3 deionized water along the cleaned glass slide through an injection pump (flow rate 6 μL / min). Restrict the movement range of the assembled film through a circular rubber tube, and increase the density of the monolayer SiO2 film by dropping 5 mM CTAB solution into the gap between the petri dish and the circular rubber tube. The finally formed film is blue-violet. When a white precipitate forms at the contact area between the glass slide and the water surface, the assembly ends. The device diagram is shown in Figure 1 。

[0071] Step 4: Transfer of the monolayer SiO2 microsphere film

[0072] Slowly lift the pretreated silicon wafer from the bottom of the film to completely transfer the film onto the silicon wafer surface. Lean it vertically against the filter membrane, and after natural drying, a large-area and uniform monolayer SiO2 microsphere mask is formed. The scanning electron microscope image is shown in Figure 2 , it can be seen that the method of the present invention can form a large-area and uniformly distributed monolayer SiO2 microsphere self-assembly template, which is beneficial to the preparation of large-area structures.

[0073] Step 5: Etching of the silicon pyramid structure

[0074] Place the silicon wafer covered with the SiO2 film in a reactive ion etching (RIE) equipment, introduce SF6 at 35 sccm, He at 35 sccm, and O2 at 5 sccm, with a radio frequency power of 50% (i.e., 50% of the maximum rated power of 150 W), and etch for 200 s to form a large-area periodic moth-eye biomimetic array structure. After etching, ultrasonically clean for 15 min, dry with nitrogen, and seal for storage.

[0075] Example 2

[0076] This example provides a method for preparing a moth-eye biomimetic nanoscale array. The specific method is the same as that of Example 1, except that the etching time in Step 5 is 300 s.

[0077] Comparative Example 1

[0078] This comparative example provides a method for preparing a nanoscale array. The specific method is the same as that in Example 1, except that the etching time in step 5 is 400 s.

[0079] Comparative Example 2

[0080] This comparative example provides a method for preparing a nanoscale array. The specific method is the same as that in Example 1, except that the etching time in step 5 is 500 s.

[0081] Scanning electron microscope images of the nanoscale arrays prepared with different etching times in Examples 1-2 and Comparative Examples 1-2 are shown in Figure 3 , and it can be seen that as the etching time increases, the nanostructures formed by etching gradually transition from smooth hexagonal arrays to sharp nanocone arrays. The structure formed by etching for 200 s has a flat surface and uniform arrangement, which can provide more attachment sites and is beneficial for the attachment of gold nanorods to the structure. When the etching time is 300 s, the etching makes the structure steeper and the surface roughness increases, which is not conducive to the attachment of gold nanorods. At the same time, due to the presence of nanocaps in the array, it is difficult for gold nanorods to enter the interior of the structure. When the etching time increases to 400 s and 500 s, the nanocones are significantly sharpened, making it difficult for gold nanorods to attach. Therefore, the etching time of 200 s is the optimal etching condition.

[0082] Example 3

[0083] This example provides a method for preparing a gold nanorod array, and the steps are as follows:

[0084] Step 1: Preparation of gold nanorod solution

[0085] Mix 0.1 mL of 25 mM HAuCl4 solution with 5 mL of 0.2 M CTAB aqueous solution, quickly inject 0.6 mL of 0.01 M NaBH4 solution, stir vigorously at 1200 rpm for 2 min, and age at room temperature for 30 min to obtain a gold nanorod seed solution;

[0086] Dissolve 7.0 g of CTAB and 1.234 g of sodium oleate in 250 mL of deionized water at 65 °C, stir to dissolve and then cool to 30 °C. Add 18 mL of 4 mM AgNO3 solution, let stand for 15 min, then add 250 mL of 1 mM HAuCl4 solution, stir at 700 rpm for 90 min until colorless. Add 1.5 mL of concentrated HCl (37 wt%) to adjust the pH to 1.5, stir at 400 rpm for 15 min, then inject 1.25 mL of 0.064 M ascorbic acid, stir vigorously at 1200 rpm for 30 s, add 800 μL of gold nanorod seed solution, stir at 900 rpm for 30 s, and then let stand for 12 h to obtain a gold nanorod suspension. The morphology of the gold nanorods is shown in Figure 4, it can be seen that the prepared gold nanorods have uniform sizes.

[0087] Step 2: Purification of gold nanorods

[0088] Take 10 mL of the gold nanorod suspension, centrifuge at 8000 rpm for 15 min, and remove the supernatant. Add 10 mL of sodium polystyrene sulfonate (Na-PSS) solution (concentration: 0.05%, mass concentration, the same below) and incubate for 1 h. Then centrifuge at 10000 rpm for 15 min, remove the supernatant, and re-add the Na-PSS solution. Repeat the washing step 3 times (the centrifugation step is repeated 3 times). Then use trisodium citrate solution (concentration: 4 mM) for washing, centrifuge at 9000 rpm for 10 min, remove the supernatant, re-add the trisodium citrate solution, and repeat the washing step 2 times. Finally, concentrate the solution by a factor of 10 to obtain the gold nanorod solution.

[0089] Step 3: Liquid-liquid interface self-assembly

[0090] Mix 1 mL of the gold nanorod solution with 1 mL of dichloromethane solution in a Teflon centrifuge tube. After shaking vigorously for 1 min, let it stand to form an ordered gold nanorod film at the water-air interface. Tilt the centrifuge tube, then slowly insert the substrate closely against the inner wall of the centrifuge tube so that the gold film covers the substrate completely. Then slowly lift it up and finally transfer it onto a silicon wafer. After it dries naturally, judge the self-assembly effect by scanning electron microscopy.

[0091] Example 4

[0092] This example provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration factor in Step 2 is 2 times.

[0093] Example 5

[0094] This example provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration factor in Step 2 is 20 times.

[0095] Example 6

[0096] This example provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of the trisodium citrate solution is 2 mM.

[0097] Example 7

[0098] This example provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of the trisodium citrate solution is 10 mM.

[0099] Example 8

[0100] This embodiment provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of the trisodium citrate solution is 20 mM.

[0101] Example 9

[0102] This embodiment provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of Na-PSS is 0.1%, and the concentration of the trisodium citrate solution is 10 mM.

[0103] Example 10

[0104] This embodiment provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of Na-PSS is 0.2%, and the concentration of the trisodium citrate solution is 10 mM.

[0105] Example 11

[0106] This embodiment provides a method for preparing a gold nanorod array. The specific method is the same as that in Example 3, except that the concentration of Na-PSS is 0.4%, and the concentration of the trisodium citrate solution is 10 mM.

[0107] In Examples 3 to 5, the concentration of Na-PSS was 0.05%, the concentration of citrate was 4 mM, and the SEM results of the gold nanorod arrays prepared with different concentration multiples of the gold nanosuspension are as Figure 5 shown. In Examples 6 to 8, the concentration multiple of the gold nanorod suspension was 10 times, the concentration of Na-PSS was 0.05% (mass concentration), and the SEM results of the gold nanorod arrays prepared with different concentrations of the trisodium citrate solution are as Figure 6 shown. In Examples 9 to 11, the concentration multiple of the gold nanorod suspension was 10 times, the concentration of citrate was 10 mM, and the SEM results of the gold nanorod arrays prepared with different concentrations of Na-PSS are as Figure 7 shown. It can be seen that when the self-assembly parameters of the gold nanorods are regulated to be that the concentration of Na-PSS is 0.2% (mass concentration), the concentration of trisodium citrate is 10 mM, and the concentration multiple is 10 times, the self-assembly conditions are optimal, and the gold nanorods can form a large-area, neat, and closely arranged array on the silicon wafer. Moreover, the large-area gold nanorod array structure can significantly improve the uniformity, sensitivity, and repeatability of the SERS substrate signal prepared.

[0108] Example 12

[0109] This embodiment provides a method for preparing a surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography. The specific steps are as follows:

[0110] Step 1: Cleaning of SiO2 microspheres and preparation of a monolayer template

[0111] Take 200 μL of the SiO2 microsphere suspension with a particle size of 500 nm, centrifuge it at 16000 rpm for 15 min, discard the supernatant, disperse the precipitate in 300 μL of ethanol, and ultrasonicate for 30 min until completely dispersed. Repeat the washing process 3 times to remove impurities.

[0112] Immerse a 1 cm × 1 cm silicon wafer in piranha solution (concentrated sulfuric acid: hydrogen peroxide = 2:1, volume ratio), and treat it at 120 °C for 50 min to remove surface organic matter and oxides.

[0113] Inject the SiO2 microsphere suspension into a petri dish containing 2 / 3 deionized water through a syringe pump (flow rate 6 μL / min), and form a monolayer SiO2 film by self-assembly at the liquid-gas interface. Slowly lift the pretreated silicon wafer from the bottom of the film to completely transfer the film onto the silicon wafer surface, and let it dry naturally for later use.

[0114] Step 2: Etching of silicon pyramid structure

[0115] Place the silicon wafer covered with the SiO2 film in a reactive ion etching (RIE) device, introduce SF6 at 35 sccm, He at 35 sccm, and O2 at 5 sccm, with a radio frequency power of 50% (i.e., 50% of the maximum rated power of 150 W), and etch for 200 s to form a periodic moth-eye biomimetic array structure. After etching, ultrasonically clean for 15 min, dry with nitrogen, and seal for storage.

[0116] Step 3: Preparation of gold nanorod solution

[0117] Mix 0.1 mL of 25 mM HAuCl4 solution with 5 mL of 0.2 M CTAB solution, quickly inject 0.6 mL of 0.01 M NaBH4 solution, stir vigorously at 1200 rpm for 2 min, and age at room temperature for 30 min to obtain a gold nanorod seed solution.

[0118] Dissolve 7.0 g of CTAB and 1.234 g of sodium oleate in 250 mL of deionized water at 65 °C, stir until dissolved, and then cool to 30 °C. Add 18 mL of 4 mM AgNO3 solution, let it stand for 15 min, then add 250 mL of 1 mM HAuCl4 solution, stir at 700 rpm for 90 min until colorless. Add 1.5 mL of concentrated HCl (37 wt%) to adjust the pH to 1.5, stir at 400 rpm for 15 min, then inject 1.25 mL of 0.064 M ascorbic acid, stir vigorously at 1200 rpm for 30 s, add 800 μL of the gold nanorod seed solution, stir at 900 rpm for 30 s, and then let it stand for 12 h to obtain a gold nanorod suspension.

[0119] Step 4: Purification and interfacial self-assembly of gold nanorods

[0120] Take 10 mL of the gold nanorod suspension, centrifuge it at 8000 rpm for 15 min, incubate it with 10 mL of 0.2 wt% sodium polystyrene sulfonate (Na-PSS) solution for 1 h first, then centrifuge and wash it twice with 10 mM sodium citrate solution, and finally concentrate it 10 times to obtain the concentrated solution.

[0121] Mix 1 mL of the concentrated solution with 1 mL of dichloromethane in a Teflon centrifuge tube, shake it vigorously for 1 min and then let it stand. An orderly arranged gold nanorod film is formed at the water-air interface.

[0122] Step 5: Fabrication of the composite substrate

[0123] Transfer the gold nanorod film obtained in Step 4 onto the moth-eye bionic array structure obtained in Step 2 (the transfer method is the same as before: tilt the centrifuge tube, then slowly insert the substrate close to the centrifuge tube wall so that the gold nanorod film covers the substrate, and then slowly lift it, and finally transfer it onto the substrate. The gold nanorod film transferred onto the substrate will further optimize the arrangement of the gold nanorods under the action of capillary force). The number of layers of the gold nanorod film is 1 layer (1layer+moth-eye), and at the same time, assemble it on a silicon wafer under the same conditions for comparison (1layer+wafer).

[0124] Example 13

[0125] This example provides a preparation method of a surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography. The specific steps are the same as those in Example 12, except that the number of layers of the gold nanorod film in Step 5 is 2 layers (2layer+moth-eye); and at the same time, assemble it on a silicon wafer under the same conditions for comparison (2layer+wafer).

[0126] Example 14

[0127] This example provides a preparation method of a surface-enhanced Raman substrate based on the self-assembly of gold nanorods and nanosphere lithography. The specific steps are the same as those in Example 12, except that the number of layers of the gold nanorod film in Step 5 is 4 layers (4layer+moth-eye); and at the same time, assemble it on a silicon wafer under the same conditions for comparison (4layer+wafer).

[0128] Scanning electron micrographs of the surface-enhanced Raman substrates based on the self-assembly of gold nanorods and nanosphere lithography with different numbers of self-assembled gold nanorod films on the silicon wafer and the moth-eye bionic array structure in Examples 12 to 14 are shown in Figure 8 , and it can be seen that as the number of self-assembled layers increases, the aspect ratio of the array structure gradually decreases, and the gold nanorods are prone to agglomeration.

[0129] Raman signal detection

[0130] The substrates prepared in Examples 12 to 14 were placed in a rhodamine (R6G) solution (10 -6 ~10 -12 M) and soaked for 30 min. The substrates adsorbed with trace amounts of R6G were placed in a Raman spectrometer for testing. The test parameters were: light source wavelength 532 nm, laser power 0.05%, integration time 10 s, and integration times 1 time. The results are shown in Figure 9 , and it can be seen that as the number of self-assembled layers increases, the performance of the substrate increases. At the same time, due to the presence of the nanoarray structure, it has better light absorption performance compared with the silicon wafer. The increase in the number of self-assembled layers provides more "hot spots" for the substrate, further increasing the SERS performance of the substrate.

[0131] In the present invention, by optimizing the content of Na-PSS, the concentration of trisodium citrate, and the concentration multiple, a large-area, neat, and closely arranged gold nanorod array structure was prepared. Compared with Patent CN119187587A, the gold nanorod array structure prepared by the method of the present invention can form a self-assembled array of gold nanorods on a centimeter scale and is applicable to large-scale production and practical applications (for example, as shown in Figure 14 , the large-area distribution of self-assembled gold nanorods on the substrate in Example 10). The gold nanorod array structure formed in Patent CN119187587A is only about 1 μm, making it difficult to achieve large-area preparation.

[0132] In the present invention, the array structure formed by reactive ion etching is combined with the self-assembly of gold nanorods, significantly improving the stability of the substrate. The stability differences between the SERS substrate prepared in the present invention (taking the composite substrate prepared in Example 12 as an example) and the single self-assembly structure (i.e., directly self-assembling gold nanorods on a silicon wafer) were compared through soaking experiments and dropping experiments. In the soaking experiment, the prepared SERS substrate and the single self-assembly structure were respectively soaked in a petri dish filled with pure water for 30 min, taken out after soaking, dried at 60 °C, and their structural changes were observed. In the dropping experiment, 20 μL of pure water was vertically dropped onto the SERS substrate and the single self-assembly structure, and after the water droplets dried naturally, their structural changes were observed. The results of the dropping experiment found that the gold nanorods on the silicon wafer were prone to stacking and irregular aggregation due to the action of liquid surface tension, resulting in the destruction of the original self-assembly structure. On the composite substrate prepared in the present invention, although a small amount of gold nanorods were lost, most of the gold nanorods still remained evenly arranged (as shown in Figure 10As shown). In addition, in the immersion experiment, due to surface tension, the gold nanorod film on the silicon wafer is easily separated from the silicon wafer and then enters the liquid environment, resulting in a large loss of gold nanorods. On the moth-eye array structure, the microstructure of the array increases the surface contact area and dispersive force. At the same time, the array structure provides more adsorption sites for the gold nanorods, enabling them to effectively resist the influence of liquid surface tension, and most of the gold nanorods remain in the array structure (as Figure 10 shown).

[0133] The SERS substrate with 1 layer of assembled layers in Example 12 was used for testing:

[0134] The prepared surface-enhanced Raman substrate was placed in a rhodamine (R6G) solution (10 -6 ~10 -12 M) and soaked for 30 min. The surface-enhanced Raman substrate adsorbed with trace amounts of R6G was placed in a Raman spectrometer for testing. The test parameters were: light source wavelength 532 nm, laser power 0.05% (0.05% of 50 mW), integration time 10 s, and integration times 1 time. The results are shown in Figure 11 , and the linear fitting results are shown in Figure 12 . It can be seen that the substrate can clearly detect a 10 -12 M concentration of R6G solution, proving that the substrate has high sensitivity and also proving that the substrate can be used for quantitative detection.

[0135] On a R6G substrate with a concentration of 10 -6 M, 20 points were randomly selected for Raman testing. The test parameters were: light source wavelength 532 nm, laser power 0.05% (0.05% of 50 mW), integration time 10 s, and integration times 1 time. The results are shown in Figure 13 . It can be seen that the SERS substrate has good uniformity and repeatability.

[0136] The above is only a preferred specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily thought of by those skilled in the art within the technical scope disclosed by the present invention should be covered by the protection scope of the present invention. Therefore, the protection scope of the present invention should be subject to the protection scope of the claims.

Claims

1. A method for preparing a surface enhanced Raman substrate based on gold nanorod self-assembly and nanosphere lithography, characterized in that: The following steps are involved: A single-layer SiO2 microsphere film is prepared by Langmuir-Blodgett method, the single-layer SiO2 microsphere film is transferred to the surface of a silicon wafer, and a moth-eye bionic nanoarray is formed by reactive ion etching, wherein the reactive ions are a mixture of SF6, He and O2; A suspension containing gold nanorods is prepared by a seed growth method, and hexadecyltrimethylammonium bromide and sodium oleate are added during the preparation of the suspension containing gold nanorods to form a gold nanorod film through liquid-liquid interface self-assembly; The gold nanorod film is transferred to the surface of the moth-eye bionic nanoarray to obtain the surface enhanced Raman substrate based on gold nanorod self-assembly and nanosphere lithography.

2. The preparation method according to claim 1, characterized in that: The steps of preparing a single-layer SiO2 film by the Langmuir-Blodgett method are: centrifuging a SiO2 microsphere suspension, discarding the supernatant, dispersing the precipitate in ethanol, and ultrasonicating until it is completely dispersed to obtain a SiO2 microsphere suspension with impurities removed; and assembling a single-layer SiO2 microsphere film on the surface of a glass slide using the SiO2 microsphere suspension with impurities removed.

3. The preparation method according to claim 1, characterized in that: Before transferring the single-layer SiO2 microsphere film to the surface of the silicon wafer, the method further includes immersing the silicon wafer in a piranha solution and pre-treating the silicon wafer under heating conditions.

4. The preparation method according to claim 1, characterized in that: The steps of forming the moth-eye bionic nanoarray by reactive ion etching are as follows: placing the silicon wafer covered with the single-layer SiO2 microsphere film in a reactive ion etching device, introducing a mixed gas of SF6, He and O2, with a radio frequency power of 50% and an etching time of 200 to 300 seconds.

5. The preparation method according to claim 1, characterized in that: The preparation steps of the gold nanorod film are as follows: a. The HAuCl4 solution was mixed with the cetyltrimethylammonium bromide solution, and the NaBH4 solution was added, stirred and aged at room temperature to obtain a gold nanorod seed solution; b. Mix cetyltrimethylammonium bromide and sodium oleate in water, add AgNO3 solution, add HAuCl4 solution after standing, stir until colorless, adjust pH to 1 to 2, add ascorbic acid, stir well, add the gold nanorod seed solution, and stand to obtain a gold nanorod suspension; c. The gold nanorod suspension is centrifuged, incubated with a sodium polystyrene sulfonate solution, then centrifuged and washed with a trisodium citrate solution, and concentrated to obtain a concentrate; d. The concentrated solution is mixed with dichloromethane in a centrifuge tube, shaken and then allowed to stand to form an orderly arranged gold nanorod film at the water-air interface.

6. The preparation method according to claim 5, characterized in that: In step b, the mass ratio of hexadecyltrimethylammonium bromide to sodium oleate is (5-7):

1.

7. The preparation method according to claim 5, characterized in that: In step c, the concentration of the sodium polystyrene sulfonate solution is 0.05-0.4 wt %.

8. The preparation method according to claim 5, characterized in that: In step c, the concentration of the trisodium citrate solution is 2-20 mM.

9. A surface enhanced Raman substrate based on gold nanorod self-assembly and nanosphere lithography, characterized in that: The preparation method according to any one of claims 1 to 8 comprises a moth-eye bionic nanoscale array, wherein gold nanorods exist between the moth-eye bionic nanoscale array.

10. Application of the surface enhanced Raman substrate based on gold nanorod self-assembly and nanosphere lithography as claimed in claim 9 in the field of Raman sensing.

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