Optical fiber microstructure biosensor for detecting alpha fetoprotein and preparation method thereof

Abstract

The invention belongs to the technical field of optical fiber sensing, and provides an optical fiber microstructure biosensor for detecting alpha fetoprotein and a preparation method thereof, and the method comprises the following steps: step 1, synthesizing a gold nanorod; 2, selecting a single-mode optical fiber for real-time monitoring, removing a coating layer, placing the single-mode optical fiber in a fusion splicer, drawing to obtain a micro-conical optical fiber, enabling CO2 laser to act on a conical waist area of the micro-conical optical fiber to manufacture a cone, taking out the optical fiber, and cooling and curing to obtain an optical fiber microstructure sensor; step 3, immersing a sensing area of the optical fiber microstructure sensor into NaOH, washing and drying, and immersing the sensing area into an MPTMS-ethanol solution; washing is carried out, and then the sensing area is immersed in a GNRs-PEG-COOH solution; washing and drying, immersing the sensing area in an EDC / NHS mixed solution for soaking, washing and drying, and putting the sensing area in an mAb solution for soaking; and washing and drying to obtain the optical fiber microstructure biosensor.

Description

Technical Field

[0001] The present invention belongs to the technical field of optical fiber sensing, and in particular relates to an optical fiber microstructure biosensor for detecting alpha-fetoprotein and a preparation method thereof. Background Art

[0002] Cancer is a major threat to public health and a global health concern. According to global cancer statistics, hepatocellular carcinoma (HCC) ranks among the top three in both morbidity and mortality. Most malignancies, including liver cancer, can be detected and treated early to reduce morbidity and mortality. However, most patients are discovered in the middle or late stages of the disease because early-stage HCC often has no obvious clinical symptoms, which greatly limits the effectiveness of treatment. Therefore, identifying early liver cancer markers is crucial for the management of liver cancer.

[0003] One of the many glycoproteins associated with HCC markers is alpha-fetoprotein (AFP). In healthy individuals, normal AFP concentrations are typically below 25 ng / mL. However, AFP levels are often elevated in HCC patients. Studies have shown a strong correlation between AFP levels and primary liver cancer, with elevated blood AFP levels positively correlated with tumor volume. Therefore, developing AFP detection methods is crucial for the early identification and treatment of HCC. Currently, AFP detection methods primarily include enzyme-linked immunosorbent assay (ELISA), electrochemical immunoassay, colorimetry, fluorescence, chemiluminescence, and Raman spectroscopy. However, while ELISA reagents are low-cost, they are time-consuming. Electrochemical devices offer the advantages of low cost and high sensitivity, but there is a trade-off between stability and sensitivity. Chemiluminescence offers high sensitivity but is costly and not suitable for physiological conditions.

[0004] Fiber optic sensors offer the advantages of compact structure, small size, label-free operation, and resistance to electromagnetic interference. In recent years, fiber optic sensing technology has shown great promise in the field of biochemistry. Changes in the refractive index (RI) induced by a detector attached to the optical fiber surface allow for extremely sensitive identification of target molecules. To enhance the sensitivity of fiber optic sensors, sensitization techniques have been developed, including fiber micromachining and surface functionalization with noble metal nanoparticles or low-dimensional nanomaterials.

[0005] The present invention is based on the method of coating gold nanorod material on an optical fiber microstructure as a sensitive material, and at the same time combining it with antibodies to prepare an optical fiber microstructure biosensor. The optical fiber microstructure biosensor can quickly detect the biomarker AFP, meeting clinical requirements and has broad application prospects in the detection of early liver cancer. Summary of the Invention

[0006] In order to solve the problems encountered in detecting alpha-fetoprotein, the present invention provides an optical fiber microstructure biosensor for detecting alpha-fetoprotein and a preparation method thereof based on a method of coating gold nanorod material on an optical fiber microstructure as a sensitive material and combining it with antibodies.

[0007] The purpose of the present invention can be achieved through the following technical solutions:

[0008] A first aspect of the present invention provides a method for preparing an optical fiber microstructure biosensor for detecting alpha-fetoprotein, comprising the following steps:

[0009] Step 1: Synthesis of gold nanorods

[0010] 1 mL of seed solution was added to 9 mL of growth solution and shaken for 10 seconds to obtain a mixed solution; 1 mL of the mixed solution was added to 9 mL of growth solution and shaken for 15 seconds, then all of it was added to 90 mL of growth solution and shaken for 15 seconds. The solution was allowed to stand in a water bath at room temperature overnight. After removing the supernatant, the gold nanorods at the bottom were collected and ultrasonically dispersed in 10 mL of deionized water to obtain a GNRs-CTAB solution;

[0011] Through the above technical solution, gold nanorods (GNRs) are a type of gold nanoparticle. Due to their inherent structure, they have transverse and longitudinal vibrations, which produce two plasmon resonance absorption peaks. As the aspect ratio increases, the longitudinal plasmon absorption peak shifts from the visible light region to the near-infrared region, and the absorption intensity is also affected by concentration. Therefore, coating gold nanorods on optical fiber microstructures can enhance the electric field. Because the wavelength of the incident light is more closely matched to the longitudinal plasmon wave of the gold nanorods, the appropriate high aspect ratio gold nanorods can stimulate a stronger LSPR effect.

[0012] Step 2: Preparation of optical fiber microstructure sensor

[0013] S1. Select a single-mode fiber (SMF) and connect both ends of the SMF to a patch cord using a cleaver. The input end of the patch cord is fixed to an ultra-wideband light source with a wavelength range of 1250-1650nm, and the output end is fixed to an optical spectrum analyzer with a resolution of 0.02nm for real-time monitoring.

[0014] S2. Remove the coating layer with a wire stripper and wipe it clean with alcohol. Place the stripped SMF in a fusion splicer to draw a micro-tapered optical fiber (T-SMF).

[0015] S3. Apply CO2 laser to the waist area of ​​T-SMF to produce a cone. This physical structural change is achieved by laser softening and drawing to obtain an ideal taper and smooth transition.

[0016] S4, taking out the optical fiber, cooling and solidifying it naturally, and obtaining an optical fiber microstructure sensor with a micro-conical long-period grating;

[0017] Step 3: Preparation of optical fiber microstructure biosensor

[0018] The sensing area of ​​the optical fiber microstructure sensor obtained in step 2 was immersed in 0.2M NaOH for 3.5 hours, washed with deionized water, and then dried in air for 0.5 hours to activate the hydroxyl groups on the fiber surface; the sensing area of ​​the dried optical fiber microstructure sensor was immersed in a 15% MPTMS-ethanol solution and maintained for 3 hours of cross-linking to modify the thiol group; because the optical fiber surface cannot directly form a stable bond with the antibody, the optical fiber surface was first alkylated to achieve covalent coupling of the antibody; the unbound MPTMS on the optical fiber surface was washed with ethanol, and the sensing area of ​​the optical fiber microstructure sensor was then immersed in a 2mL GNRs-PEG-COOH solution for 4 hours;

[0019] The sensing area of ​​the microstructured optical fiber sensor was cleaned with alcohol and deionized water in sequence and dried in air for 15 minutes. The sensing area of ​​the optical fiber microstructure sensor was then immersed in an EDC / NHS mixed solution for 20 minutes to activate the carboxyl functional groups on the gold nanorods. The optical fiber microstructure sensor was rinsed with phosphate buffer and dried in air. It was then immersed in a mAb solution (pH 7.4) for 0.5 hours. The amino groups (-NH2 groups) in the mAb solution gradually combined with the carboxyl groups (-COOH groups) on the sensor surface. Finally, the sensing area of ​​the sensor was cleaned with PBS buffer to remove the free mAb that failed to bind to the optical fiber surface and dried in air for 5 minutes to obtain the mTLPG-LSPR biosensor (i.e., micro-tapered long-period fiber grating localized surface plasmon biosensor, referred to as optical fiber microstructure biosensor).

[0020] Furthermore, the seed solution in step 1 is prepared by the following steps:

[0021] A chloroauric acid (HAuCl4) solution was added to a cetyltrimethylammonium bromide (CTAB) solution and stirred for 2 minutes. Then, a glacial sodium borohydride (NaBH4) solution was added, stirred for 1 minute, and allowed to stand for 1 hour to obtain a seed solution. In the cetyltrimethylammonium bromide surfactant, the chloroauric acid was rapidly reduced by the glacial sodium borohydride, forming gold nanocrystal seeds. Gold nanorods were then synthesized using a hierarchical growth method.

[0022] Furthermore, in step 1, the dosage ratio of hexadecyltrimethylammonium bromide, chloroauric acid solution and sodium borohydride solution is 9.2 mL:0.25 mL:500 μL; wherein the concentration of hexadecyltrimethylammonium bromide is 0.1 M, the concentration of chloroauric acid solution is 0.01 M, and the concentration of sodium borohydride solution is 0.01 mol / L.

[0023] Furthermore, the growth solution in step 1 is prepared by a three-step growth method.

[0024] Furthermore, the growth solution in step 1 is 2.5×10 -4 M HAuCl4, 5.0×10 -4 M ascorbic acid (AA) and 0.1M CTAB. CTAB acts as a soft template to guide the directional growth of gold nanostructures. The weak reducing agent ascorbic acid (AA) converts the gold ions (Au) in HAuCl4 into 3+ ) is gradually reduced to elemental gold (Au 0 ) and selectively deposited on the surface of the gold seed, causing the gold seed to extend in a specific direction and eventually forming a gold nanorod structure with a specific aspect ratio.

[0025] Furthermore, the micro-conical long-period grating in S4 of step 2 is a micro-conical long-period grating with a cone diameter of 20 μm, a grating period of 400 μm, and a grating length of 4 mm.

[0026] Furthermore, the EDC / NHS mixed solution in step 3 is prepared by the following steps:

[0027] 0.04 g of EDC and 0.02 g of NHS were weighed and dissolved in 2 mL of deionized water at a mass ratio of 2:1 to obtain an EDC / NHS mixed solution.

[0028] Furthermore, the mAb solution in step 3 is prepared by the following steps:

[0029] 10 μL of 1 mg / mL mAb (monoclonal antibody) was weighed and added dropwise to 10 μL of phosphate buffer to prepare a mAb solution.

[0030] Furthermore, the GNRs-PEG-COOH solution in step 3 is prepared by the following steps:

[0031] To 10 mL of the GNRs-CTAB solution obtained in step 1, add 10 mL of a 0.01 g / mL thiol polyethylene glycol carboxylic acid (HS-PEG-COOH) solution. Stir overnight, centrifuge, and ultrasonically disperse with 10 mL of deionized water for 5 minutes. By forming an Au-S covalent bond, the CTAB is replaced by the thiol ligand to obtain a carboxyl-coated gold nanorod (GNRs-PEG-COOH) solution. Because the gold nanorod surface cannot form a stable bond with the antibody directly, the gold nanorods synthesized in step 1 are carboxylated to achieve covalent coupling with the antibody.

[0032] The second aspect of the present invention provides an optical fiber microstructure biosensor for detecting alpha-fetoprotein obtained by the preparation method described in the first aspect.

[0033] Beneficial effects of the present invention:

[0034] This invention utilizes a method of coating gold nanorods onto optical fiber microstructures as a sensitive material and combining them with antibodies to create a fiber-optic microstructure biosensor. The combination of gold nanorods, antibodies, and optical fiber sensing technology leverages the advantages of all three, simplifying sensor design while improving system reliability and durability. The resulting fiber-optic microstructure biosensor is capable of rapidly detecting the biomarker AFP, meeting clinical requirements and possessing broad application prospects in early-stage liver cancer detection.

[0035] The fiber-optic microstructured biosensor provided by the present invention has an LSPR sensor with a micro-conical long-period grating (mTLPG) structure, which can be used to detect AFP, providing rapid, accurate, and targeted detection of AFP with minimal detection time and high specificity. The mTLPG structure only requires SMF for fabrication; at the same time, to improve the sensitivity of the sensor, an optimal concentration of gold nanorods (GNRs) is modified within the sensing area. The surface of the GNRs is also modified with a large number of carboxyl groups, enabling effective binding with monoclonal antibodies (mAbs), giving our sensor the advantage of a fast response time and achieving an effective detection time of approximately 3 minutes.

[0036] The present invention utilizes the combination of optical fiber microstructure and local surface plasma resonance to improve the sensitivity of the optical fiber.

[0037] The optical fiber microstructure sensor provided by the present invention directly detects the wavelength drift of the binding of alpha-fetoprotein and antibodies in the near-infrared band.

[0038] The optical fiber microstructure biosensor provided by the present invention has high sensitivity and rapid response capability: by coating gold nanorod material on the surface of the optical fiber microstructure as a sensitizer and fixing antibodies on the surface of the optical fiber microstructure, this design significantly enhances the sensitivity, response speed and selectivity of the sensor to alpha-fetoprotein.

[0039] The optical fiber microstructure biosensor provided by the present invention is resistant to electromagnetic interference, miniaturized, and lightweight: by utilizing the advantages of optical fiber sensing technology, the sensor exhibits good anti-electromagnetic interference capabilities, while having the characteristics of miniaturization and lightweight, and is suitable for application in a variety of environments.

[0040] The preparation process of the optical fiber microstructure biosensor provided by the present invention is simple: the preparation process of the optical fiber microstructure biosensor for measuring alpha-fetoprotein is relatively simple, including the synthesis of gold nanorod materials, the manufacture of microstructured optical fibers, and the coating of materials, all of which are easy to operate. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] The present invention will be further described below with reference to the accompanying drawings.

[0042] Figure 1 This is a physical picture of the gold nanorod solution prepared by the present invention at room temperature;

[0043] Figure 2 This is a physical picture of the optical fiber fusion splicer for the microstructure optical fiber sensor prepared by the present invention;

[0044] Figure 3 This is an electron microscope image of the gold nanorods prepared by the present invention on the surface of a microstructured optical fiber structure;

[0045] Figure 4 Schematic diagram of the biosensor experimental device prepared by the present invention;

[0046] Figure 5 This is a graph showing the spectrum changes of alpha-fetoprotein at different concentrations measured by the biosensor prepared by the present invention;

[0047] Figure 6 This is a linear fitting diagram of wavelength changes when the biosensor prepared by the present invention measures alpha-fetoprotein at different concentrations;

[0048] Figure 7 This is a diagram showing the wavelength signal changes of various proteins measured by the biosensor prepared by the present invention. DETAILED DESCRIPTION

[0049] The following will be combined with the embodiments to clearly and completely describe the technical solutions of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0050] The single-mode optical fiber (SMF) used in the present invention is a G652D single-mode optical fiber with a specification of 9 / 125 μm.

[0051] Example 1

[0052] This embodiment provides a fiber optic microstructure biosensor for detecting alpha-fetoprotein and a preparation method thereof:

[0053] A method for preparing an optical fiber microstructure biosensor for detecting alpha-fetoprotein comprises the following steps:

[0054] Step 1: Synthesis of gold nanorods

[0055] Prepare the seed solution: Add 0.25 mL of 0.01 M chloroauric acid (HAuCl4) solution to 9.2 mL of 0.1 M cetyltrimethylammonium bromide (CTAB) solution and stir for 2 minutes. The solution changes from colorless to light yellow and then to orange-yellow. Then, add 500 μL of 0.01 mol / L freshly prepared icy sodium borohydride (NaBH4) solution and stir for 1 minute. The solution turns brown-purple and is allowed to stand for 1 hour to obtain the seed solution.

[0056] Preparation of growth solution: The growth solution was prepared by three-step growth method. The specific growth solution was: 2.5×10 -4 MHAuCl4, 5.0×10 -4 M ascorbic acid (AA) and 0.1 M CTAB;

[0057] Synthesis of gold nanorods: 9 mL of growth solution was injected into containers A and B respectively, and 90 mL of growth solution was injected into container C; 1 mL of seed solution was added to container A containing 9 mL of growth solution, shaken for 10 seconds, and the solution turned light purple to obtain a mixed solution; then 1 mL of the mixed solution was added to container B containing 9 mL of growth solution, shaken for 15 seconds, and the solution turned light purple; finally, all the solution in container B was transferred to container C containing 90 mL of growth solution, shaken for 15 seconds, and allowed to stand in a water bath at room temperature overnight, and the solution turned purple; after removing the supernatant of container C, the gold nanorods at the bottom were collected, and 10 mL of deionized water was added for ultrasonic dispersion to obtain a GNRs-CTAB solution with a concentration of 3 mg / mL (such as Figure 1 As shown), the solution turns brown, and the gold nanorod material is obtained;

[0058] Step 2: Preparation of optical fiber microstructure sensor

[0059] S1. Select a single-mode fiber (SMF) and connect both ends of the SMF to a patch cord using a cleaver. The input end of the patch cord is fixed to an ultra-wideband light source with a wavelength range of 1250-1650nm, while the output end is fixed to an optical spectrum analyzer with a resolution of 0.02nm to achieve real-time monitoring.

[0060] S2. Use wire strippers to remove 2.5cm of coating layer and wipe it clean with alcohol cotton pad. Set the mode and parameters, adjust the motor, and place the stripped SMF on the LZM-100 welding machine (such as Figure 2 As shown), a micro-tapered optical fiber (T-SMF) is drawn;

[0061] S3. Use a motor to move the starting area of ​​the T-SMF's waist to the working area of ​​the CO2 laser, so that the CO2 laser acts on the waist area to produce a cone. This physical structural change is achieved through laser softening and drawing to obtain the ideal taper and smooth transition.

[0062] S4. Remove the optical fiber and allow it to solidify by natural cooling to ensure that the newly formed structure is stable, thereby obtaining an optical fiber microstructure sensor with a micro-conical long-period grating (conical region diameter of 20 μm, grating period of 400 μm, and grating length of 4 mm);

[0063] Step 3: Preparation of optical fiber microstructure biosensor

[0064] Weigh 0.04 g of EDC and 0.02 g of NHS at a mass ratio of 2:1 and dissolve them in 2 mL of deionized water to obtain an EDC / NHS mixed solution;

[0065] Weigh 10 μL of 1 mg / mL mAb (monoclonal antibody) and add it dropwise to 10 μL of phosphate buffer to prepare mAb solution;

[0066] To 10 mL of GNRs-CTAB solution, 10 mL of 0.01 g / mL thiol polyethylene glycol carboxylic acid (HS-PEG-COOH) solution was added, stirred overnight, centrifuged twice (8000 rpm, 10 min / time), and then ultrasonically dispersed with 10 mL of deionized water for 5 min. By forming Au-S covalent bonds, CTAB was replaced by thiol ligands to obtain a gold nanorod (GNRs-PEG-COOH) solution with carboxyl groups coated on the surface;

[0067] The sensing area of ​​the microstructured optical fiber sensor obtained in step 2 was immersed in 0.2 M NaOH for 3.5 hours, washed with deionized water, and then air-dried for 0.5 hours to activate the hydroxyl groups on the fiber surface; the sensing area of ​​the dried optical fiber microstructure sensor was immersed in a 15% MPTMS-ethanol solution and kept cross-linked for 3 hours to modify the thiol groups; the unbound MPTMS on the optical fiber surface was washed with ethanol, and the sensing area of ​​the optical fiber microstructure sensor was then immersed in a 2 mL GNRs-PEG-COOH solution for 4 hours;

[0068] The sensing area of ​​the microstructured optical fiber sensor was cleaned multiple times with an alcohol solution, then cleaned twice with deionized water and air-dried for 15 minutes. The sensing area of ​​the optical fiber sensor was immersed in an EDC / NHS mixed solution for 20 minutes to activate the carboxyl functional groups on the gold nanorods. The optical fiber sensor was rinsed with phosphate buffer and air-dried. It was then immersed in a mAb solution (pH 7.4) for 0.5 hours. The amino groups (-NH2 groups) in the mAb solution gradually combined with the carboxyl groups (-COOH groups) on the sensor surface. Finally, the sensing area of ​​the sensor was cleaned with PBS buffer to remove free mAb that failed to bind to the optical fiber surface, and air-dried for 5 minutes to obtain the mTLPG-LSPR biosensor (i.e., micro-tapered long-period fiber grating localized surface plasmon biosensor, referred to as optical fiber microstructure biosensor).

[0069] The optical fiber microstructure biosensor for detecting alpha-fetoprotein is prepared by the above steps.

[0070] The optical fiber microstructure biosensor prepared in Example 1 was applied and tested:

[0071] Application: Fix the prepared optical fiber microstructure biosensor on a highly uniform platform, connect one end to an ultra-wideband light source, and the other end to a spectrum analyzer. The experimental setup is shown in the figure below. Figure 4 shown.

[0072] Test: During the entire test process, the external temperature was maintained at room temperature. The analyte was immersed in a customized mold (half glass tube, r = 2mm, R = 4mm), ensuring that the immersion level remained consistent each time. At the beginning of the test, the light source was turned on, the entire optical path was kept stationary, and alpha-fetoprotein solutions of different concentrations were selected for immersion. As the concentration of alpha-fetoprotein increased, the central wavelength of the resonant wave detected by the spectrometer shifted; the wavelength change of the output spectrum of alpha-fetoprotein (AFP) is shown in the figure below. Figure 5-6 shown.

[0073] To further evaluate the feasibility and performance of the fiber-optic microstructured biosensor for measuring alpha-fetoprotein in practical applications, the following tests were performed:

[0074] Specificity test: Four different proteins (bovine serum albumin (BSA), carcinoembryonic antigen (CEA), prostate antigen (PSA), and prostate cancer RNA (PCA3)) were selected for control experiments. The concentration of each protein tested was fixed at 400 ng / mL. The wavelength change of the output spectrum of each protein is shown in the figure below. Figure 7 shown.

[0075] Depend on Figure 7It can be seen that the binding of bovine serum albumin (BSA), carcinoembryonic antigen (CEA), prostate antigen (PSA) and prostate cancer RNA (PCA3) is very small. The experimental results show that the mTLPG-LSPR fiber biosensor for measuring AFP has strong specificity.

[0076] It should be noted that, in this document, relational terms such as first and second, etc., are used only to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any actual relationship or order between these entities or operations. Moreover, the terms "comprises," "comprising," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that includes a list of elements includes not only those elements but also other elements not explicitly listed, or elements inherent to such process, method, article, or apparatus.

[0077] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A method for preparing an optical fiber microstructure biosensor for detecting alpha-fetoprotein, characterized in that: The following steps are involved: Step 1: Synthesis of gold nanorods 1 mL of seed solution was added to 9 mL of growth solution and shaken to obtain a mixed solution; 1 mL of the mixed solution was added to 9 mL of growth solution and shaken, and all of it was added to 90 mL of growth solution, shaken, and allowed to stand in a water bath at room temperature overnight. The gold nanorods were collected and ultrasonically dispersed in deionized water to obtain a GNRs-CTAB solution; Step 2: Preparation of optical fiber microstructure sensor S1. Select single-mode optical fiber for real-time monitoring; S2, after removing the coating layer, placing the single-mode optical fiber in a fusion splicer to draw a micro-tapered optical fiber; S3, applying a CO2 laser to the waist region of the micro-tapered optical fiber to produce a cone; S4, taking out the optical fiber, cooling and solidifying it naturally, and obtaining an optical fiber microstructure sensor with a micro-conical long-period grating; Step 3: Preparation of optical fiber microstructure biosensor The sensing area of ​​the optical fiber microstructure sensor obtained in step 2 was immersed in NaOH, washed and dried, and the sensing area was immersed in MPTMS-ethanol solution; After washing, the sensing area was immersed in the GNRs-PEG-COOH solution; Wash and dry, immerse the sensing area in an EDC / NHS mixed solution, wash and dry, and immerse in a mAb solution; wash and dry to obtain an optical fiber microstructure biosensor.

2. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The seed solution in step 1 is prepared by the following steps: Add chloroauric acid solution to the hexadecyltrimethylammonium bromide solution and stir for 2 minutes; then add icy sodium borohydride solution, stir for 1 minute, and let stand for 1 hour to obtain a seed solution.

3. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 2, characterized in that: In step 1, the dosage ratio of cetyltrimethylammonium bromide, chloroauric acid solution, and sodium borohydride solution is 9.2 mL:0.25 mL:500 μL; wherein the concentration of cetyltrimethylammonium bromide is 0.1 M, the concentration of chloroauric acid solution is 0.01 M, and the concentration of sodium borohydride solution is 0.01 mol / L.

4. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The growth solution in step 1 is prepared by a three-step growth method.

5. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The growth solution in step 1 is 2.5×10 -4 M HAuCl4, 5.0×10 -4 M ascorbic acid and 0.1 M CTAB.

6. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The micro-conical long-period grating in S4 of step 2 is a micro-conical long-period grating with a cone diameter of 20 μm, a grating period of 400 μm, and a grating length of 4 mm.

7. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The EDC / NHS mixed solution in step 3 is prepared by the following steps: 0.04 g of EDC and 0.02 g of NHS were weighed and dissolved in 2 mL of deionized water at a mass ratio of 2:1 to obtain an EDC / NHS mixed solution.

8. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: The mAb solution in step 3 was prepared by the following steps: 10 μL of 1 mg / mL mAb was weighed and added dropwise to 10 μL of phosphate buffer to prepare a mAb solution.

9. The method for preparing a fiber optic microstructure biosensor for detecting alpha-fetoprotein according to claim 1, characterized in that: In step 3, the GNRs-PEG-COOH solution was prepared by the following steps: To 10 mL of the GNRs-CTAB solution obtained in step 1, 10 mL of 0.01 g / mL mercaptopolyethylene glycol carboxylic acid solution was added, stirred overnight, centrifuged, and ultrasonically dispersed with deionized water to obtain a GNRs-PEG-COOH solution.

10. An optical fiber microstructure biosensor for detecting alpha-fetoprotein, characterized in that: The invention is prepared by the preparation method according to any one of claims 1 to 9.

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