Preparation method and application of sandwich type photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material

Abstract

The application relates to a preparation method and application of a sandwich type photoelectrochemical immunosensor based on a Bi2S3 / BiOCl composite material, and belongs to the field of novel functional materials and biosensing detection technology; the Bi2S3 / BiOCl composite material is used as a base material, and visible light is used as a light source to obtain a photoelectric current; the two components of the base material are well matched in bands, greatly improving photoelectric conversion efficiency, so that a large output signal is obtained; different concentrations of the NSE solution to be measured result in different amounts of immune complexes, further resulting in different degrees of influence on photoelectric signals; the sensor shows high sensitivity, a wide detection range and a low detection limit to target objects, can realize rapid and sensitive determination of NSE in human serum, and has great application value for judging diseases of patients in the clinic.

Description

TECHNICAL FIELD

[0001] The application relates to a preparation method and application of a photoelectrochemical immunosensor based on a Bi2S3 / BiOCl composite material, in particular to a sandwich type photoelectrochemical immunosensor for sensitively detecting NSE by adopting a Bi2S3 / BiOCl composite material as a base material, and belongs to the technical field of novel functional materials and biosensing detection. BACKGROUND

[0002] The photoelectric conversion process of a photoelectrochemical (PEC) immunosensor is realized by a photoelectric active material. Under the excitation of light, electrons jump from a valence band to a conduction band, and an electron-hole pair is generated. The separation of the electron and the hole generates a photovoltage, and a photocurrent is formed in an external circuit, so that a light signal is converted into an electric signal. Based on this principle, it is crucial to seek a photoelectric active material with high conversion performance, high stability and non-toxicity for constructing a photoelectrochemical sensor. At present, TiO2 is most studied. Because of a wide band gap, TiO2 can only respond to ultraviolet light (<400 nm) which accounts for only 4% of the total energy of sunlight. Developing a visible light responsive photoelectric material is one of the most important ways to realize high-efficiency photoelectric conversion.

[0003] BiOCl is a typical V-VI-VII ternary semiconductor compound, and has many unique properties, such as a characteristic layered structure, sufficient chemical stability, a tunable band gap (3.3 eV) and superior photocatalytic activity. However, the application of BiOCl in photoelectrochemical sensors is limited due to its small specific surface, relatively positive conduction band position, weak reduction power, poor dispersion, unsatisfactory visible light absorption and short lifetime of photoinduced electron-hole pairs. The band gap of Bi2S3 is very narrow (1.3 ~ 1.7 eV), which can well match the band gap of BiOCl, so that good photoelectric performance is obtained, the utilization rate of light is improved, and the excitation of the light signal is enhanced.

[0004] In the preparation process of the PEC immunosensor, the transfer of electrons is hindered due to steric hindrance. Therefore, most PEC immunosensors belong to the signal attenuation type. A covalent metal framework (COF) can be used as a good signal tag for connecting a detection antibody Ab2 to amplify the signal because the COF has the advantages of large surface area, good biocompatibility, high stability, low cost, poor conductivity and easy modification. The Ag-NH2 bond between a silver nanoparticle (Ag NP) and an antibody can effectively increase the number of Ab2 molecules on the surface of the composite material. The Ag NPs can be formed in situ on the surface of the COF material to obtain TpPa@CeO2-Ag NPs.

[0005] Different levels of NSE are fixed on the electrode through specific binding of antigen and antibody, and then a fixed concentration of TpPa@CeO2-Ag NPs-Ab2 is combined on the electrode through specific reaction of NSE and Ab2, so that the photocurrent is obviously reduced; the sandwich type PEC immunosensor has ultrahigh sensitivity, good reproducibility and acceptable specificity. SUMMARY

[0006] One of the purposes of the present application is to obtain Bi2S3 material with superior performance by a hydrothermal method, and further optimize photoelectric conversion activity by modification such as compounding;

[0007] The second purpose of the present application is to prepare a sandwich type photoelectrochemical immunosensor with high sensitivity, strong specificity and fast detection speed based on a Bi2S3 / BiOCl composite material as a substrate, so as to realize ultrahigh sensitive detection of NSE under visible light.

[0008] The technical scheme of the present application is as follows:

[0009] 1. A preparation method of a sandwich type photoelectrochemical immunosensor based on a Bi2S3 / BiOCl composite material, characterized in that it comprises the following steps:

[0010] 1) Cut ITO conductive glass into 1.6*0.6 cm2, and sequentially ultrasonic clean with detergent powder, acetone, ethanol and ultrapure water for 0.5 h, and dry under nitrogen;

[0011] 2) Add 6 µL of 1 ~ 6 mg / mL Bi2S3 / BiOCl solution to the conductive surface of the ITO conductive glass, and dry at room temperature;

[0012] 3) Add 1 ~ 5 mg / mL neuron-specific enolase (NSE) Ab1 solution to the surface of the modified electrode, and incubate at 4 DEG C for 1 h, then clean the electrode surface with ultrapure water, and naturally air dry to a wet film state

[0013] 4) Add 6 µL of 1% ~ 3% bovine serum albumin solution to the surface of the modified electrode, and after 0.5 h, rinse the electrode surface with ultrapure water, and air dry to a wet film state in a 4 DEG C refrigerator;

[0014] 5) Add 6 µL of NSE standard solution with different concentrations, and after 4 h of constant temperature incubation at 4 DEG C, rinse the electrode surface with ultrapure water;

[0015] 6) Add 6 µL of COFs-silver nanoparticle conjugated NSE antibody (TpPa@CeO2-Ag NPs-Ab2) standard solution, and after 4 h of constant temperature incubation at 4 DEG C, rinse the electrode surface with ultrapure water, to obtain a working electrode.

[0016] 2. The NSE antibody solution with a concentration of 1 pg / mL according to claim 1 is obtained by diluting a 1 mg / mL NSE antibody solution purchased from Nanjing Kingsriver Biotech Co., Ltd. with a phosphate buffer.

[0017] 3. The TpPa@CeO2-Ag NPs-Ab2 solution with a fixed concentration according to claim 1 is obtained by dissolving 3 ~ 6 mg TpPa@CeO2-Ag NPs in 1 mL PBS (pH = 7.4), then adding a 1 mg / mL NSE antibody standard solution purchased from Nanjing Kingsriver Biotech Co., Ltd., and diluting with a phosphate buffer.

[0018] 4. The phosphate buffer according to claim 1 is prepared by mixing a 0.1 mol / L sodium phosphate dibasic solution with a 0.1 mol / L potassium phosphate monobasic solution, and adjusting the pH to 7.4.

[0019] 5. The TpPa@CeO2-Ag NPs solution according to claim 3 is obtained by gradually adding 1 wt% ~ 5 wt% ammonia water to a 10 mg / mL AgNO3 solution to obtain a silver ammine solution as a Ag NPs precursor solution; and dissolving 50 mg ~ 80 mg TpPa@CeO2 powder in 40 mL or more of solution, and continuously stirring at room temperature in the dark.

[0020] 6. The TpPa@CeO2 powder according to claim 5 is prepared by adding 0.1 ~ 0.3 mol NaOH and 10 ~ 35 mg TpPa to 35 mL deionized water, stirring for 10 min, then dissolving 2 mmol Ce(NO3)3·6H2O in the above solution, stirring for 30 min, heating at 100 ℃ for 24 h, cooling to room temperature, centrifugal separation, washing with deionized water and ethanol, and vacuum drying at 60 ℃.

[0021] 7. The TpPa according to claim 6 is prepared by dissolving 2,4,6-trihydroxybenzene-1,3,5-tricarboxaldehyde (TP) and p-phenylenediamine (Pa) in a molar ratio of 4:9 in a mixed solution of 1.5 mL 1,4-dioxane and 1.5 mL trimethylbenzene, ultrasonicating for 15 min, slowly adding 0.5 mL of a 3 mol / L acetic acid solution; reacting at 120 ℃ for 3 days; centrifugal separation, washing with acetone, and vacuum drying at 60 ℃ for 12 h.

[0022] 8. The Bi2S3 / BiOCl solution according to claim 1, wherein the preparation steps are as follows:

[0023] Bismuth nitrate pentahydrate and thiourea were dissolved in 60 mL of deionized water at a molar ratio of 1:2, stirred and dissolved, and then the mixture was transferred to a reaction kettle for reaction at 160 DEG C for 12 h; after natural cooling, the product was collected, washed with ultrapure water and ethanol by centrifugation, and dried at 60 DEG C under vacuum for 12 h, and after grinding, Bi2S3 powder was obtained;

[0024] Bismuth nitrate pentahydrate, sodium chloride and mannitol were added to 40 mL of deionized water at a molar ratio of 1:1:2, stirred and dissolved, and the synthesized Bi2S3 powder was added, and stirring was continued for 30 min; reaction was carried out at 160 DEG C for 12 h, and after natural cooling, the product was collected, washed with ultrapure water and ethanol, and dried at 60 DEG C under vacuum overnight to obtain Bi2S3 / BiOCl powder; the Bi2S3 / BiOCl powder was dispersed in deionized water to obtain a Bi2S3 / BiOCl solution.

[0025] 9. A sandwich type photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material prepared by the preparation method of claim 1 for detecting NSE, characterized in that the detection step is as follows:

[0026] 1) An electrochemical workstation was used to test in a three-electrode system, a saturated calomel electrode was used as a reference electrode, a platinum wire electrode was used as an auxiliary electrode, and a modified ITO electrode was used as a working electrode, and the test was carried out in 15 mL of a phosphate buffer solution with a concentration of 0.05 ~ 0.25 mol / L of ascorbic acid at pH 5.0 ~ 8.0;

[0027] 2) The NSE standard solution was detected by time-current method, the voltage was set to 0 V, the running time was 120 s, and the light source wavelength was 400 ~ 500 nm;

[0028] 3) After the electrode was placed, the light was turned on every 20 s for 20 s, the photocurrent was recorded, and the working curve was drawn;

[0029] 4) The NSE sample solution to be detected was used instead of the NSE standard solution for detection.

[0030] Advantages of the present application

[0031] 1. The present application first applies BiOCl material to construct a PEC sensing model, and solves the problem of low photoelectric conversion efficiency of the material alone by the compounding of Bi2S3, and the composite material has great application potential in photoelectric conversion;

[0032] 2. The sandwich photoelectrochemical immunosensor prepared by the method can be used for NSE detection, has short response time, low detection limit, wide linear range and good stability, can realize simple, rapid, high-sensitivity and specific detection, and provides a novel analysis method for early clinical detection of NSE. Specific embodiments

[0033] Example 1: A preparation method of a sandwich photoelectrochemical immunosensor based on a Bi2S3 / BiOCl composite material:

[0034] 1) ITO conductive glass was cut into 1.6*0.6 cm 2 , and sequentially cleaned with decontamination powder, acetone, ethanol and ultrapure water for 0.5 h, and dried under nitrogen blowing;

[0035] 2) 6 μL of a Bi2S3 / BiOCl solution with a concentration of 1 mg / mL was added to the conductive surface of the ITO conductive glass, and dried at room temperature;

[0036] 3) 5 mg / mL of a neuron-specific enolase (NSE) Ab1 solution was added to the surface of the modified electrode, and incubated at 4°C for 1 h, then cleaned with ultrapure water, and naturally air-dried to a wet film state

[0037] 4) 6 μL of a 2% bovine serum albumin solution was added to the surface of the modified electrode, and the electrode surface was cleaned with ultrapure water after 0.5 h, and air-dried to a wet film state in a 4°C refrigerator;

[0038] 5) 6 μL of an NSE standard solution with different concentrations was added, and the electrode surface was cleaned with ultrapure water after 4 h of constant temperature incubation at 4°C;

[0039] 6) 6 μL of a COFs-silver nanoparticle conjugated NSE antibody (TpPa@CeO2-Ag NPs-Ab2) standard solution was added, and the electrode surface was cleaned with ultrapure water after 4 h of constant temperature incubation at 4°C, to obtain a working electrode.

[0040] Example 2: A preparation method of a sandwich photoelectrochemical immunosensor based on a Bi2S3 / BiOCl composite material:

[0041] 1) ITO conductive glass was cut into 1.6*0.6 cm 2 , and sequentially cleaned with decontamination powder, acetone, ethanol and ultrapure water for 0.5 h, and dried under nitrogen blowing;

[0042] 2) 6 μL of a Bi2S3 / BiOCl solution with a concentration of 3 mg / mL was added to the conductive surface of the ITO conductive glass, and dried at room temperature;

[0043] 3) Drop 3 mg / mL neuron-specific enolase (NSE) Ab1 solution on the surface of the modified electrode, incubate at 4°C for 1 h, then wash with ultrapure water, and naturally air dry to a wet film state

[0044] 4) Drop 6 μL of 1% bovine serum albumin solution on the surface of the modified electrode, wash the electrode surface with ultrapure water after 0.5 h, and air dry to a wet film state in a 4°C refrigerator;

[0045] 5) Drop 6 μL of NSE standard solution at different concentration levels, wash the electrode surface with ultrapure water after 4 h of constant temperature incubation at 4°C;

[0046] 6) Drop 6 μL of COFs-silver nanoparticle conjugated NSE antibody (TpPa@CeO2-Ag NPs-Ab2) standard solution, wash the electrode surface with ultrapure water after 4 h of constant temperature incubation at 4°C, and prepare the working electrode.

[0047] Example 3: Preparation method of a sandwich type photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material:

[0048] 1) Cut the ITO conductive glass into 1.6 x 0.6 cm 2 , ultrasonically clean with detergent powder, acetone, ethanol and ultrapure water for 0.5 h, and dry under nitrogen;

[0049] 2) Drop 6 μL of 6 mg / mL Bi2S3 / BiOCl solution on the conductive surface of the ITO conductive glass, and air dry at room temperature;

[0050] 3) Drop 1 mg / mL neuron-specific enolase (NSE) Ab1 solution on the surface of the modified electrode, incubate at 4°C for 1 h, then wash with ultrapure water, and naturally air dry to a wet film state

[0051] 4) Drop 6 μL of 3% bovine serum albumin solution on the surface of the modified electrode, wash the electrode surface with ultrapure water after 0.5 h, and air dry to a wet film state in a 4°C refrigerator;

[0052] 5) Drop 6 μL of NSE standard solution at different concentration levels, wash the electrode surface with ultrapure water after 4 h of constant temperature incubation at 4°C;

[0053] 6) Drop 6 μL of COFs-silver nanoparticle conjugated NSE antibody (TpPa@CeO2-Ag NPs-Ab2) standard solution, wash the electrode surface with ultrapure water after 4 h of constant temperature incubation at 4°C, and prepare the working electrode.

[0054] Example 4: Preparation of Bi2S3 / BiOCl composite material:

[0055] Dissolve 6 mmol of bismuth nitrate pentahydrate and 12 mmol of thiourea in 60 mL of deionized water, stir to dissolve, and then transfer the mixture to a reaction kettle for reaction at 160 °C for 12 h; after natural cooling, collect the product, wash with ultrapure water and ethanol, centrifugalize, and dry at 60 °C under vacuum for 12 h; after grinding, obtain Bi2S3 powder;

[0056] Add 2 mmol of bismuth nitrate pentahydrate, 2 mmol of sodium chloride, and 4 mmol of mannitol to 40 mL of deionized water, stir to dissolve, add the synthesized Bi2S3 powder, and continue stirring for 30 min; react at 160 °C for 12 h, after natural cooling, collect the product, wash with ultrapure water and ethanol, centrifugalize, and dry at 60 °C under vacuum overnight to obtain Bi2S3 / BiOCl powder; grind the powder to prepare an aqueous solution with a concentration of 3 mg / mL.

[0057] Example 5 Preparation of Bi2S3 / BiOCl composite material:

[0058] Dissolve 20 mmol of bismuth nitrate pentahydrate and 40 mmol of thiourea in 60 mL of deionized water, stir to dissolve, and then transfer the mixture to a reaction kettle for reaction at 160 °C for 12 h; after natural cooling, collect the product, wash with ultrapure water and ethanol, centrifugalize, and dry at 60 °C under vacuum for 12 h; after grinding, obtain Bi2S3 powder;

[0059] Add 1 mmol of bismuth nitrate pentahydrate, 1 mmol of sodium chloride, and 2 mmol of mannitol to 40 mL of deionized water, stir to dissolve, add the synthesized Bi2S3 powder, and continue stirring for 30 min; react at 160 °C for 12 h, after natural cooling, collect the product, wash with ultrapure water and ethanol, centrifugalize, and dry at 60 °C under vacuum overnight to obtain Bi2S3 / BiOCl powder; grind the powder to prepare an aqueous solution with a concentration of 1 mg / mL.

[0060] Example 6 Preparation of Bi2S3 / BiOCl composite material:

[0061] Dissolve 12 mmol of bismuth nitrate pentahydrate and 24 mmol of thiourea in 60 mL of deionized water, stir to dissolve, and then transfer the mixture to a reaction kettle for reaction at 160 °C for 12 h; after natural cooling, collect the product, wash with ultrapure water and ethanol, centrifugalize, and dry at 60 °C under vacuum for 12 h; after grinding, obtain Bi2S3 powder;

[0062] Bismuth nitrate pentahydrate (0.5 mmol), sodium chloride (0.5 mmol) and mannitol (0.5 mmol) were added to 40 mL of deionized water, stirred to dissolve, and Bi2S3 powder synthesized above was added, and stirring was continued for 30 min; the reaction was carried out at 160 °C for 12 h, and after natural cooling, the product was collected, washed with ultrapure water and ethanol, and dried at 60 °C under vacuum overnight to obtain Bi2S3 / BiOCl powder; the powder was ground and prepared into an aqueous solution with a concentration of 6 mg / mL.

[0063] Example 7 Preparation of TpPa@CeO2-Ag NPs-Ab2 solution:

[0064] TpPa was prepared by dissolving 63.0 mg of 2,4,6-trihydroxybenzene-1,3,5-tricarboxaldehyde (TP) and 48.0 mg of p-phenylenediamine (Pa) in a mixed solution of 1.5 mL of 1,4-dioxane and 1.5 mL of trimethylbenzene, and ultrasonicating for 15 min, and then slowly adding 0.5 mL of acetic acid with a concentration of 3 mol / L; the reaction was carried out at 120 °C for 3 days; the product was separated by centrifugation, washed with acetone, and dried at 60 °C under vacuum for 12 h;

[0065] TpPa@CeO2 powder was prepared by adding 7 g of NaOH and 20 mg of TpPa to 35 mL of deionized water, stirring for 10 min, then dissolving 2 mmol of Ce(NO3)3·6H2O in the above solution, stirring for 30 min, heating at 100 °C for 24 h, cooling to room temperature, and then separating by centrifugation, washing with deionized water and ethanol, and drying at 60 °C under vacuum;

[0066] 1 wt% ammonia water was gradually added to a 10 mg / mL AgNO3 solution to obtain a silver ammonia solution as a Ag NPs precursor solution; 80 mg of TpPa@CeO2 powder was dissolved in 40 mL of the above solution, and the resulting solution was continuously stirred at room temperature in the dark;

[0067] 3 mg of TpPa@CeO2-Ag NPs was dissolved in 1 mL of PBS (pH = 7.4), and then 1 mg / mL of NSE antibody standard solution was added and diluted with phosphate buffer.

[0068] Example 8 Preparation of TpPa@CeO2-Ag NPs-Ab2 solution:

[0069] TpPa was prepared by dissolving 94.5 mg of 2,4,6-trihydroxybenzene-1,3,5-tricarboxaldehyde (TP) and 72 mg of p-phenylenediamine (Pa) in a mixed solution of 1.5 mL of 1,4-dioxane and 1.5 mL of trimethylbenzene, ultrasonic for 15 min, slowly add 0.5 mL of acetic acid with a concentration of 3 mol / L; react for 3 days at 120 °C; centrifugal separation, wash with acetone, and vacuum dry at 60 °C for 12 h.

[0070] TpPa@CeO2 powder was prepared by adding 12 g of NaOH and 10 mg of TpPa to 35 mL of deionized water, stirring for 10 min, then dissolving 2 mmol of Ce(NO3)3·6H2O in the above solution, stirring for 30 min, heating at 100 °C for 24 h, cooling to room temperature, centrifugal separation, washing with deionized water and ethanol, and vacuum drying at 60 °C.

[0071] 3 wt% ammonia water was gradually added to a 10 mg / mL AgNO3 solution to obtain a silver ammonia solution as a Ag NPs precursor solution; 50 mg of TpPa@CeO2 powder was dissolved in 40 mL of the above solution, and the resulting solution was continuously stirred at room temperature in the dark;

[0072] 6 mg of TpPa@CeO2-Ag NPs was dissolved in 1 mL of PBS (pH = 7.4), then 1 mg / mL of NSE antibody standard solution was added, and diluted with phosphate buffer.

[0073] Example 9 Preparation of SiO2 / PDA-Ag NPs-Ab2 solution:

[0074] TpPa was prepared by dissolving 31.5 mg of 2,4,6-trihydroxybenzene-1,3,5-tricarboxaldehyde (TP) and 24 mg of p-phenylenediamine (Pa) in a mixed solution of 1.5 mL of 1,4-dioxane and 1.5 mL of trimethylbenzene, ultrasonic for 15 min, slowly add 0.5 mL of acetic acid with a concentration of 3 mol / L; react for 3 days at 120 °C; centrifugal separation, wash with acetone, and vacuum dry at 60 °C for 12 h.

[0075] TpPa@CeO2 powder was prepared by adding 10 g of NaOH and 10 mg of TpPa to 35 mL of deionized water, stirring for 10 min, then dissolving 2 mmol of Ce(NO3)3·6H2O in the above solution, stirring for 30 min, heating at 100 °C for 24 h, cooling to room temperature, centrifugal separation, washing with deionized water and ethanol, and vacuum drying at 60 °C.

[0076] After gradually adding 2 wt% of ammonia water into 10 mg / mL AgNO3 solution, a silver ammonia solution was obtained as a precursor solution of Ag NPs; 60 mg of TpPa@CeO2 powder was dissolved in 40 mL of the above solution, and the resulting solution was continuously stirred at room temperature in the dark.

[0077] 6 mg of TpPa@CeO2-Ag NPs was dissolved in 1 mL of PBS (pH = 7.4), and then 1 mg / mL of NSE antibody standard solution was added and diluted with phosphate buffer.

[0078] Example 10: Detection of NSE:

[0079] (1) The electrochemical workstation was used to test in a three-electrode system, with a saturated calomel electrode as the reference electrode, a platinum wire electrode as the auxiliary electrode, and the prepared ITO modified electrode as the working electrode. The test was performed in 15 mL of PBS buffer solution with a concentration of 0.1 mol / L ascorbic acid and a pH of 5.5;

[0080] (2) The NSE standard solution was detected by time-current method, with a voltage of 0 V, a running time of 120 s, and a light source wavelength of 400 nm;

[0081] (3) After the electrode was placed, the light was turned on every 20 s for 20 s, and the photocurrent was recorded to draw the working curve;

[0082] (4) The NSE sample solution was used instead of the NSE standard solution for detection.

[0083] Example 11: Detection of NSE:

[0084] (1) The electrochemical workstation was used to test in a three-electrode system, with a saturated calomel electrode as the reference electrode, a platinum wire electrode as the auxiliary electrode, and the prepared ITO modified electrode as the working electrode. The test was performed in 15 mL of PBS buffer solution with a concentration of 0.2 mol / L ascorbic acid and a pH of 7.4;

[0085] (2) The NSE standard solution was detected by time-current method, with a voltage of 0 V, a running time of 120 s, and a light source wavelength of 450 nm;

[0086] (3) After the electrode was placed, the light was turned on every 20 s for 20 s, and the photocurrent was recorded to draw the working curve;

[0087] (4) The NSE sample solution was used instead of the NSE standard solution for detection.

[0088] Example 12: Detection of NSE:

[0089] (1) The test was carried out using an electrochemical workstation in a three-electrode system, with a saturated calomel electrode as a reference electrode, a platinum wire electrode as an auxiliary electrode, and the prepared ITO modified electrode as a working electrode, in a 15 mL PBS buffer solution with a concentration of 1.5 mol / L ascorbic acid and pH of 8.0;

[0090] (2) The NSE standard solution was detected by time-current method, the voltage was set to 0 V, the running time was 120 s, and the light source wavelength was 500 nm;

[0091] (3) After the electrode was placed, the light was turned on every 20 s for 20 s, the photocurrent was recorded, and the working curve was drawn;

[0092] (4) The NSE sample solution to be detected was used instead of the NSE standard solution for detection.

Claims

1. A preparation method of a sandwich-type photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material, characterized in that, The method comprises the following steps: 1) Cut ITO conductive glass into 1.6 x 0.6 cm 2 , and sequentially clean with decontamination powder, acetone, ethanol, and ultrapure water for 0.5 h, and dry under nitrogen. 2) 6 μL of Bi2S3 / BiOCl solution with a concentration of 1-6 mg / mL is added dropwise to the conductive surface of ITO conductive glass, and is dried at room temperature; 3) 1-5 mg / mL of neuron-specific enolase Ab1 solution is added dropwise to the surface of the modified electrode, and after incubation at 4 DEG C for 1 h, the electrode surface is washed with ultrapure water and naturally air-dried to a wet film state; 4) 6 μL of 1%-3% bovine serum albumin solution is added dropwise to the surface of the modified electrode, and after 0.5 h, the electrode surface is washed with ultrapure water and air-dried to a wet film state in a refrigerator at 4 DEG C; 5) 6 μL of NSE standard solution with different concentrations is added dropwise, and after incubation at 4 DEG C for 4 h, the electrode surface is washed with ultrapure water; 6) 6 μL of COFs-silver nanoparticle conjugated NSE antibody standard solution, namely TpPa@CeO2-AgNPs-Ab2 solution, is added dropwise, and after incubation at 4 DEG C for 4 h, the electrode surface is washed with ultrapure water to obtain a working electrode; The preparation method of the COFs-silver nanoparticle conjugated NSE antibody standard solution, namely TpPa@CeO2-AgNPs-Ab2 solution, is as follows: TpPa is prepared by dissolving 2,4,6-trihydroxybenzene-1,3,5-triformaldehyde and p-phenylenediamine in a mixed solution of 1.5 mL of 1,4-dioxane and 1.5 mL of trimethyltoluene at a molar ratio of 4:9, ultrasonicating for 15 min, slowly adding 0.5 mL of 3 mol / L acetic acid, reacting at 120 DEG C for 3 days, centrifuging, washing with acetone, and vacuum drying at 60 DEG C for 12 h; TpPa@CeO2 powder is prepared by adding 0.1-0.3 mol of NaOH and 10-35 mg of TpPa to 35 mL of deionized water, stirring for 10 min, then dissolving 2 mmol of Ce(NO3)3.6H2O in the above solution, stirring for 30 min, heating at 100 DEG C for 24 h, cooling to room temperature, centrifuging, washing with deionized water and ethanol, and vacuum drying at 60 DEG C to obtain the powder; 1-5 wt% of ammonia water is gradually added to a 10 mg / mL AgNO3 solution to obtain a silver ammonia solution as a Ag NPs precursor solution; 80 mg of TpPa@CeO2 powder is dissolved in 40 mL of the above solution, and the TpPa@CeO2-Ag NPs solution is prepared by continuously stirring at room temperature in the dark; 3 mg of TpPa@CeO2-AgNPs is dissolved in 1 mL of pH 7.4 phosphate buffer solution, then 1 mg / mL of NSE antibody standard solution is added, and the TpPa@CeO2-AgNPs-Ab2 solution is obtained by dilution with a phosphate buffer.

2. The method for preparing the sandwich photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material according to claim 1, characterized in that, The Bi2S3 / BiOCl solution is prepared by the following steps: 1) Bismuth nitrate pentahydrate and thiourea are dissolved in 60 mL of deionized water at a molar ratio of 1:2, and then the mixture is transferred to a reaction kettle for reaction at 160 DEG C for 12 h; after natural cooling, the product is collected, washed with ultrapure water and ethanol by centrifugation, and vacuum dried at 60 DEG C for 12 h to obtain Bi2S3 powder; 2) Bi(NO3)3.5H2O, NaCl and mannitol were added into 40 mL deionized water with the molar ratio of 1:1:2, stirred and dissolved, and the synthesized Bi2S3 powder was added, and stirring was continued for 30 min; the reaction was carried out at 160℃ for 12 h, and after natural cooling, the product was collected, washed with ultrapure water and ethanol, and dried at 60℃ under vacuum overnight to obtain Bi2S3 / BiOCl powder; the Bi2S3 / BiOCl powder was dispersed in deionized water to obtain a Bi2S3 / BiOCl solution.

3. The sandwich type photoelectrochemical immunosensor based on Bi2S3 / BiOCl composite material prepared by the preparation method of claim 1, characterized in that, The sensor is used for neuron-specific enolase detection, and the detection steps are as follows: 1) The electrochemical workstation is used to test in a three-electrode system, a saturated calomel electrode is used as a reference electrode, a platinum wire electrode is used as an auxiliary electrode, and a sandwich-type photoelectrochemical immunosensor is used as a working electrode, and the test is carried out in 15 mL of a phosphate buffer solution with a pH of 5.0-8.0 and a concentration of 0.05-0.25 mol / L of ascorbic acid; 2) The NSE standard solution is detected by time-current method, the voltage is set to 0 V, the running time is 120 s, and the light source wavelength is 400-500 nm; 3) After the electrode is placed, the light is turned on every 20 s for 20 s, the photocurrent is recorded, and the working curve is drawn; 4) The NSE sample solution to be detected is used to replace the NSE standard solution for detection.