Preparation method and application of ternary heterojunction composite photoelectrochemical immunosensor

Abstract

The application relates to a preparation method and application of a ternary heterojunction composite photoelectrochemical immunosensor, and belongs to the technical field of photoelectrochemical biosensors, and particularly relates to a preparation method and application of a photoelectrochemical immunosensor for detecting alpha-fetoprotein by adopting a BiPO4 / BiOBr / CdS composite material. The method is characterized in that a ternary heterojunction is assembled on an FTO modified electrode to fix a capture antibody, and the accurate determination of alpha-fetoprotein is realized through a photocurrent signal response. The BiPO4 / BiOBr heterojunction expands the forbidden band width and expands the light absorption range; due to the sensitization of CDs, the absorption of visible light is enhanced, so that the photocurrent signal is increased. The method has the advantages of good stability, strong specificity, high sensitivity, good reproducibility and the like, is good for the rapid detection of alpha-fetoprotein, solves the existing technical difficulties, and provides a feasible method for the detection of tumor markers.

Description

Technical Field

[0001] This invention belongs to the field of photoelectrochemical biosensor technology, specifically relating to a method for preparing a BiPO4 / BiOBr / CdS ternary heterojunction composite photoelectrochemical immunosensor and its application in the detection of alpha-fetoprotein. Background Technology

[0002] In recent years, cancer has become one of the most common diseases and a major public health problem. Although the mortality rate of cancer is high, early diagnosis and treatment intervention can significantly improve the cure rate of cancer patients. Alpha-fetoprotein (AFP) is a glycoprotein belonging to the albumin family, mainly synthesized by fetal hepatocytes and the yolk sac. AFP concentration is high in fetal blood circulation, gradually decreasing after birth. By 2-3 months after birth, AFP is largely replaced by albumin, making it difficult to detect in the blood, resulting in extremely low levels in adult serum. AFP has many important physiological functions, including transport functions, bidirectional regulatory functions as a growth regulator, immunosuppression, and induction of T lymphocyte apoptosis. AFP is closely related to the occurrence and development of liver cancer and various tumors, showing high concentrations in various tumors and serving as a positive indicator for tumor detection. Currently, it is mainly used clinically as a serum marker for primary liver cancer, for diagnosis and monitoring of treatment efficacy. Generally, the concentration of AFP in human serum is less than 20 ng / mL. -1 However, levels exceeding this threshold may indicate hepatocellular carcinoma. As an important biomarker, early, sensitive, and rapid monitoring of abnormal changes in AFP levels is crucial for the early diagnosis of primary liver cancer.

[0003] As a promising molecular detection and analysis technology, photoelectrochemical chromatography (PEC) combines the advantages of both photochemistry and electrochemistry, namely high sensitivity, fast response, low background response, and simple operation, making it highly valuable for protein detection, DNA detection, and biomarker detection. In photoelectrochemical detection, when a photoactive material is excited by a light source, photogenerated electrons and holes separate and transfer or undergo redox reactions, generating a photocurrent as a detection signal. Over the past few decades, methods such as ratiometric fluorescence, aptamer-based fluorescence, enzyme-linked immunosorbent assay (ELISA), inductively coupled plasma mass spectrometry (ICP-MS), electrochemical immunoassay, and electrochemiluminescence have been developed as effective methods for detecting AFP. However, these existing methods suffer from drawbacks such as high equipment costs, time consumption, and complex procedures.

[0004] As the core of photoelectric sensors, photosensitive materials have a significant impact on their performance. BiPO4, a bismuth metal semiconductor, is often used as a photoactive material to participate in photochemical reactions due to its non-toxic and corrosion-resistant properties. However, BiPO4's wide bandgap (4.40 eV) results in a weak response to visible light, and its high electron-hole recombination rate limits its application in photoelectrochemical sensors. Coupled with two photoelectric materials to form a heterojunction is considered an effective way to solve this problem, significantly improving the photoelectric conversion efficiency of the composite material. Generally, BiPO4 and BiOBr are commonly used to construct photocatalytic materials, but there are few reports on using BiPO4 and BiOBr heterojunctions to establish photoelectrochemical sensors. Cadmium sulfide (CdS) has strong light absorption in the visible light region and is an extremely important photoelectric material. Due to its narrow bandgap, CdS is considered an excellent material for PEC analysis and can be used in efficient photosensitive BiPO4 / BiOBr heterojunctions to respond to visible light, further improving the material's photoelectric response to visible light.

[0005] This invention couples two optoelectronic materials, BiPO4 and BiOBr, to form a heterojunction, and then further uses CdS quantum dots for sensitization. The matched band structure greatly improves the photoelectric efficiency of the sensor. Summary of the Invention

[0006] To address the problems existing in the prior art, this invention provides a method for preparing a BiPO4 / BiOBr / CdS ternary heterojunction composite photoelectrochemical immunosensor and its application in the detection of alpha-fetoprotein. The photoelectrochemical sensor constructed in this invention utilizes the principle of specific binding between antigen and antibody for detection. The preparation method is simple and the structure is stable. Furthermore, this method is harmless to humans and the environment, and has the advantages of high specificity, high accuracy, simple operation, and faster processing, which is helpful for rapid trace detection.

[0007] The present invention solves the above-mentioned technical problems by adopting the following solution:

[0008] This invention provides a method for preparing a ternary heterojunction composite photoelectrochemical immunosensor, wherein the heterojunction is a BiPO4 / BiOBr / CdS ternary heterojunction composite material, comprising the following steps:

[0009] Step 1: Preparation of BiPO4 / BiOBr composite material;

[0010] Under stirring conditions, NaBr, NaH2PO4, and Bi(NO3)3·5H2O were added to a solvent and mixed vigorously to obtain a mixed solution. The mixed product was ultrasonicated for 10-20 min, then stirred for another 20-40 min at room temperature. The mixture was then transferred to a polytetrafluoroethylene-lined reactor and heated at 160-220℃ for 12-24 h. After the reaction was complete, the mixture was allowed to cool naturally, and the precipitate was collected by centrifugation. The precipitate was washed 2-4 times with anhydrous ethanol and ultrapure water, respectively. The precipitate was then dried at 40-80℃ for 12-24 h to obtain BiPO4 / BiOBr heterojunction powder.

[0011] The BiPO4 / BiOBr composite material prepared by the above method has advantages such as good light absorption, narrow band gap, no pollution, harmlessness to humans and the environment, simple synthesis, and easy availability of materials. Ultimately, BiPO4 / BiOBr composite materials with a mass percentage of 10-50 wt% can be synthesized, with a preferred BiPO4 / BiOBr heterojunction weight ratio of 40 wt%, which can obtain a stronger photocurrent.

[0012] Step 2: Preparation of water-soluble CdS QDs;

[0013] CdCl2 was added to deionized water to prepare a CdCl2 solution. After adding mercaptopropionic acid, the solution was degassed with nitrogen for 10-40 min, and then the pH was adjusted to 9-12 with NaOH solution. Na2S solution was added, and the mixture was heated under reflux at 100-120℃ for 2-6 h. After the solution cooled to room temperature, isopropanol was added for precipitation. The solid precipitate was obtained by centrifugation, washed 2-3 times with anhydrous ethanol, and dried in a vacuum drying oven at 40-80℃ for 10-24 h to obtain CdS QDs powder.

[0014] Step 3: Preparation of BiPO4 / BiOBr / CdS ternary heterojunction composite material;

[0015] BiPO4 / BiOBr heterojunction powder and CdS QDs powder were weighed separately, added to deionized water, and vortexed for 5-10 min to obtain a mixed solution. The solution was then sonicated for 30-90 min to bond the two materials, resulting in a CdS-sensitized BiPO4 / BiOBr composite suspension, which was stored at 4℃ in the dark for later use.

[0016] Step 4: Preparation of the PEC immunosensor;

[0017] Clean the FTO glass electrode, uniformly drop a CdS-sensitized BiPO4 / BiOBr / CdS suspension onto the glass electrode, dry it, and then drop AFP antibody, BSA, and AFP antigen of different concentrations onto the dried glass electrode.

[0018] The PEC immunosensor prepared by the method of this invention improves the photocurrent response through CdS sensitization, enabling effective detection of alpha-fetoprotein (AFP). It offers advantages such as high detection sensitivity, good adsorption selectivity, rapid detection, stable performance, and strong signal response. Furthermore, the preparation method utilizes widely available and inexpensive raw materials that are safe and non-toxic, and the preparation process is simple and easy to operate.

[0019] In step 1, the molar ratio of NaBr, NaH2PO4 and Bi(NO3)3·5H2O is (0.5-0.9):(0.5-0.1):1.

[0020] In step 1, the mixed product is heated in a polytetrafluoroethylene-lined reactor, preferably at 180°C for 24 hours. Heating at 180°C for 12-24 hours results in a higher yield of the synthesized composite material, where BiPO4 and BiOBr can combine to form a heterojunction and exhibit good stability.

[0021] In step 1, the preferred drying temperature and time is 60℃ for 12 hours.

[0022] In step 2, the molar ratio of CdCl2 to Na2S in the mixed solution is 1:(0.5-1), and the material ratio of CdCl2 to mercaptopropionic acid is 100-500 μl of mercaptopropionic acid per 0.001 mol of CdCl2. The concentration of the Na2S solution is 0.1 mol·L⁻¹. -1 .

[0023] In step 2, mercaptopropionic acid is added as a crosslinking agent, which facilitates the bonding of positively charged cadmium atoms and negatively charged sulfur atoms. Before the reaction, nitrogen is used for degassing to completely remove oxygen from the reaction system, eliminating oxygen interference and allowing the reaction to proceed smoothly.

[0024] In step 2, the pH value is preferably adjusted to 11 using sodium hydroxide solution. Since mercaptopropionic acid is acidic, and the reaction system needs to be carried out under alkaline conditions, the reaction is more complete when the pH is between 9 and 12, and optimally when the pH is adjusted to 11. Strictly controlling the pH value of the reaction system and heating under reflux at 100-120°C helps in the synthesis of CdS quantum dots.

[0025] In step 3, the mass ratio of BiPO4 / BiOBr heterojunction powder to CdS QDs powder is 1:1-1:5, preferably 1:3. Strictly controlling the mass ratio of BiPO4 / BiOBr heterojunction powder to CdS QDs powder can result in a strong photocurrent effect. When the mass ratio is between 1:1 and 1:5, the photocurrent first increases and then decreases. When the mass ratio is 1:3, the photocurrent reaches its maximum value; further increases in the mass ratio do not significantly enhance the photocurrent intensity.

[0026] In step 4, a 0.6×2.0cm material is used. 2 The FTO glass electrode was cleaned using acetone and 1.0 mol·L⁻¹, respectively. -1 The FTO glass electrode was ultrasonically cleaned for 10-20 min with a mixture of NaOH and 50% ethanol (v / v, 1:1) and ultrapure water. After cleaning, the FTO glass electrode was dried at 40-80℃ for 2-6 h. Ultrasonic cleaning in acetone facilitates the removal of lipid-soluble impurities, while cleaning in the NaOH and ethanol / water mixture is for modification of the FTO glass electrode. Cleaning in ultrapure water helps remove residual acetone or NaOH from the previous two steps.

[0027] In step 4, the concentration is 1.0-5.0 mg·mL -1 A CdS-sensitized BiPO4 / BiOBr / CdS suspension was uniformly dropwise onto the FTO surface and then dried at 40-80℃ for 10-24 h. Choosing an appropriate suspension concentration is crucial for the sensor's properties. When the suspension concentration is low, the material's photocurrent response is weak, resulting in low sensor sensitivity; conversely, when the suspension concentration is high, the incubated composite material exceeds the glass electrode's load limit and easily detaches from the glass electrode, leading to poor stability of the PEC sensor. Preferably, the suspension concentration is 3.0 mg·mL⁻¹. -1 .

[0028] In step 4, preferably, 10-50 μl is prepared at a concentration of 5-20 μg·mL. -1 AFP antibody was dropwise added to the surface of the BiPO4 / BiOBr / CdS modified electrode. Then, 10-50 μl of 0.5-2 wt% BSA solution was added dropwise to the surface of the modified electrode. Finally, 10-50 μl of 0.001-1000 ng·mL⁻¹ solution was added. -1 AFP antigen was uniformly dropped onto the surface of the modified electrode to modify it with different concentrations, thereby generating photocurrents of varying intensities. After each modification, the modified electrode was thoroughly washed with PBS buffer to remove excess free protein from each step, and then incubated at 37°C (the optimal temperature for proteins, which is conducive to the binding of biomolecules). Further optimization yielded a concentration of 10 μg / mL in 30 μl solutions. -1 The AFP antibody was chosen because it facilitates drying, and 30 μl is sufficient to cover the modified electrode surface. This concentration was selected to conserve antibody solution while obtaining accurate results. Further preferred, 30 μl of 1 wt% BSA was added dropwise to the surface of the modified electrode and incubated for 90 min to block non-specific binding sites. If the concentration is too high, BSA is difficult to dissolve in aqueous solution; if the concentration is too low, it will not effectively block non-specific binding sites.

[0029] Another objective of this invention is to provide an application of a ternary heterojunction composite photoelectrochemical immunosensor in the detection of alpha-fetoprotein, specifically comprising the following steps:

[0030] (1) A three-electrode system was used: the prepared PEC immunosensor was used as the working electrode; the Ag / AgCl electrode was used as the reference electrode; and the platinum electrode was used as the counter electrode. The supernatant of the human serum sample was taken as the initial sample after centrifugation three times and stored in a refrigerator at 4°C.

[0031] (2) At a concentration of 0.1 mol·L -1 Ascorbic acid (AA) was added to the PBS buffer solution as an electron donor.

[0032] (3) The excitation light source is a 250W xenon lamp, which is switched on and off every 20 seconds, with an external voltage of 0.0V.

[0033] (4) The photocurrent curves of the working electrode under illumination and non-illumination conditions were collected using an electrochemical workstation. The xenon lamp was switched on and off once every 20 seconds as one cycle, and the difference between the highest and lowest points on the vertical axis was taken as the photocurrent intensity.

[0034] (5) Plot the standard curve about AFP concentration: Plot the standard curve about AFP concentration with the photocurrent intensity of the sensor as the vertical axis and the AFP concentration as the horizontal axis to obtain the standard curve graph.

[0035] (6) Detection of AFP concentration in unknown solution: The unknown solution is incubated on the prepared PEC immunosensor, the photocurrent intensity of the sensor is detected, and the concentration of AFP in the unknown solution can be obtained by combining the standard curve.

[0036] The Ag / AgCl electrode, used as a reference electrode, ensured the reproducibility of the experiment. The platinum electrode, used as the counter electrode, ensured unimpeded current flow on the working electrode, guaranteeing that the studied reaction occurred on the working electrode. The addition of ascorbic acid (AA) to the PBS buffer solution, acting as an electron donor, facilitated the timely consumption of electron and hole vacancies on the electrode during the reaction, promoting its continuation. Adopting these parameters resulted in better photocurrent performance.

[0037] The beneficial effects of this invention are:

[0038] A PEC immunosensor based on a ternary heterojunction composite material was successfully constructed for the first time, used for the quantitative detection of alpha-fetoprotein (AFP) in serum samples. Its advantages are as follows: First, compared with pure BiPO4, the BiPO4 / BiOBr heterojunction has a narrower band gap, expanding the light absorption range and allowing for full utilization of light energy. Second, the separation of photogenerated electron-hole pairs during PEC detection is due to the effective energy level matching between BiPO4, BiOBr, and CdS, which suppresses the recombination of photogenerated electrons, thereby improving the performance of the PEC immunosensor. Third, the presence of CdS QDs enhances the sensor's light absorption capacity, generating more electron-hole pairs and increasing the photocurrent signal. Under optimal conditions, the linear range of the PEC immunosensor for AFP is 0.001-1000 ng / mL. -1 The detection limit is 0.82 pg·mL. -1 Compared with traditional sensors, it has a wider detection range, higher sensitivity, and good reproducibility, stability, and high sensitivity. Attached Figure Description

[0039] Figure 1 To investigate the photocurrent under different BiPO4 to BiOBr ratios;

[0040] Figure 2 To investigate the photocurrent under different BiPO4 / BiOBr and CdS ratios;

[0041] Figure 3 To investigate the photocurrent of BiPO4 / BiOBr / CdS suspensions with different concentrations;

[0042] Figure 4 To investigate the photocurrent under different concentrations of AA;

[0043] Figure 5 To investigate the effects of different antibody incubation times;

[0044] Figure 6 To investigate the effects of different antigen incubation times;

[0045] Figure 7 The results show the sensor photocurrent intensity under different AFP concentrations;

[0046] Figure 8 The results show the specificity comparison of the PEC bioimmunosensor prepared by this method.

[0047] Figure 9 The repeatability comparison results of the PEC bioimmunosensor prepared by this method;

[0048] Figure 10 The stability comparison results of the PEC bioimmunosensor prepared by this method are shown. Detailed Implementation

[0049] The principles and features of the present invention are described below with reference to the accompanying drawings. The embodiments described are only for explaining the present invention and are not intended to limit the scope of the present invention.

[0050] This invention provides a method for preparing a BiPO4 / BiOBr / CdS ternary heterojunction composite photoelectrochemical immunosensor, the specific steps of which are as follows:

[0051] Step 1: Preparation of BiPO4 / BiOBr composite material;

[0052] BiPO4 / BiOBr composite materials were synthesized using a one-step hydrothermal method.

[0053] Under stirring conditions, NaBr, NaH2PO4, and Bi(NO3)3·5H2O were added to methanol in a molar ratio of (0.5-0.9):(0.5-0.1):1 and stirred vigorously for 10-15 min. The mixture was ultrasonically treated for 10-20 min, then stirred again at room temperature for 20-40 min. The mixture was then transferred to a polytetrafluoroethylene-lined reactor and heated at 160-220 °C for 12-24 h. After the reactor cooled naturally, the precipitate was collected by centrifugation, and the product was washed 2-3 times with anhydrous ethanol and ultrapure water. Finally, the precipitate was dried in a forced-air drying oven at 40-80 °C for 12-24 h to obtain BiPO4 / BiOBr heterojunction powder.

[0054] Step 2: Preparation of water-soluble CdS quantum dots;

[0055] CdCl2 solution was prepared by adding CdCl2 to deionized water. After adding mercaptopropionic acid, the solution was degassed with nitrogen for 10–40 min, and then dissolved in 1.0 mol·L⁻¹ water. -1 Adjust the pH to 9-12 with NaOH solution. Finally, add 0.1 mol·L⁻¹ NaOH solution to the mixed solution. - 1 The Na₂S solution was heated under reflux at 100-120℃ for 2-6 hours. After cooling to room temperature, isopropanol was added for precipitation, and the solid precipitate was obtained by centrifugation. The precipitate was washed 2-3 times with anhydrous ethanol and dried in a vacuum drying oven at 40-80℃ for 10-24 hours to obtain CdS QDs powder. The molar ratio of CdCl₂ to Na₂S in the above mixed solution was 1:(0.5-1), and the material ratio of CdCl₂ to mercaptopropionic acid was 100-500 μl of mercaptopropionic acid per 0.001 mol CdCl₂.

[0056] Step 3: Preparation of BiPO4 / BiOBr / CdS composite material;

[0057] Water-soluble CdS quantum dots were loaded onto a BiPO4 / BiOBr heterojunction using a physical bonding method.

[0058] First, weigh out BiPO4 / BiOBr heterojunction powder and CdS QDs powder according to a mass ratio of 1:1-1:5. Add deionized water to dissolve the mixture and vortex for 5-10 minutes to ensure thorough mixing. Then, sonicate for 30-90 minutes to bond the two materials, yielding a concentration of 1.0-5.0 mg / mL. -1 The CdS-sensitized BiPO4 / BiOBr composite suspension was stored in a 4°C refrigerator away from light for later use.

[0059] Step 4: Preparation of the PEC immunosensor;

[0060] Take 0.6 × 2.0 cm 2 The FTO glass electrode was prepared with acetone and 1.0 mol·L⁻¹. -1 The FTO glass electrode was ultrasonically cleaned for 10-20 min with a mixture of NaOH and 50% ethanol (v / v, 1:1) and ultrapure water. After cleaning, the FTO glass electrode was dried at 40-80℃ for 2-6 h. A concentration of 1.0-5.0 mg / mL was prepared. -1 The CdS-sensitized BiPO4 / BiOBr composite suspension was uniformly added dropwise to FTO and dried in a drying oven at 40-80℃ for 10-24 h. The composite electrode was then successfully prepared. Then, 10-50 μl of a 5-20 μg / mL solution was added dropwise. -1 AFP antibody was prepared and incubated at 37°C to modify the surface of a BiPO4 / BiOBr / CdS modified electrode. Next, 10-50 μl of BSA (0.5-2 wt%) was added dropwise to the surface of the modified electrode to block non-specific binding sites. Finally, 10-50 μl of a solution with a concentration of 0.001-1000 ng / mL was added. -1 The AFP antigen was uniformly dropped onto the surface of the modified electrode and incubated to generate specific immune binding. The sensor was prepared at room temperature, and the PEC immunosensor was stored at 4°C until testing. To remove excess free protein, the modified electrode was thoroughly washed with PBS buffer after each modification.

[0061] The application of the BiPO4 / BiOBr / CdS ternary heterojunction composite photoelectrochemical immunosensor in the detection of alpha-fetoprotein (AFP) is as follows: The specific operation is as follows:

[0062] (1) A three-electrode system was used: the prepared PEC immunosensor was used as the working electrode; the Ag / AgCl electrode was used as the reference electrode; and the platinum electrode was used as the counter electrode. The supernatant of the human serum sample was taken as the initial sample after centrifugation three times and stored in a refrigerator at 4°C.

[0063] (2) At a concentration of 0.1 mol·L -1 Ascorbic acid (AA) was added to the PBS buffer solution as an electron donor.

[0064] (3) The excitation light source is a 250W xenon lamp, which is switched on and off every 20 seconds, with an external voltage of 0.0V.

[0065] (4) The photocurrent curves of the working electrode under illumination and non-illumination conditions were collected using an electrochemical workstation. The xenon lamp was switched on and off once every 20 seconds as one cycle, and the difference between the highest and lowest points on the vertical axis was taken as the photocurrent intensity.

[0066] (5) Plot the standard curve about AFP concentration: Plot the standard curve about AFP concentration with the photocurrent intensity of the sensor as the vertical axis and the AFP concentration as the horizontal axis to obtain the standard curve graph.

[0067] (6) Detection of AFP concentration in unknown solution: The unknown solution is incubated on the prepared PEC immunosensor, the photocurrent intensity of the sensor is detected, and the concentration of AFP in the unknown solution can be obtained by combining the standard curve.

[0068] In the following examples, the antibody (Ab) and antigen (Ag) of AFP, and carcinoembryonic antigen (CEA) were purchased from Shanghai Zeye Biotechnology Co., Ltd. (Shanghai, China). Bi(NO3)3·5H2O, ascorbic acid (AA), chromium chloride (CdCl2), and sodium sulfide (Na2S) were purchased from Shanghai McLean Biochemical Technology Co., Ltd. (Shanghai, China). Mercaptopropionic acid (MPA) was purchased from Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). Sodium dihydrogen phosphate (NaH2PO4) and sodium bromide (NaBr) were purchased from Tianjin Hengxing Chemical Reagent Manufacturing Co., Ltd. (Tianjin, China).

[0069] Unless otherwise specified, all experimental materials used in this invention can be prepared by conventional methods in the field or obtained through commercial purchases.

[0070] I. Investigation of the ratio of BiPO4 to BiOBr

[0071] Example 1

[0072] Under stirring conditions, NaH₂PO₄, NaBr, and Bi(NO₃)₃·5H₂O were simultaneously mixed in methanol and stirred vigorously for 15 min, with a molar ratio of 1:9:10. The mixture was ultrasonicated for 15 min, then stirred at room temperature for another 30 min, and then transferred to a polytetrafluoroethylene-lined reactor and heated at 180 °C for 24 h. After the reactor cooled naturally, the precipitate was collected by centrifugation, and the product was washed three times with anhydrous ethanol and ultrapure water. Finally, the precipitate was dried in a forced-air drying oven for 12 h to obtain 10 wt% BiPO₄ / BiOBr heterojunction powder for subsequent experiments.

[0073] Example 2 differs from Example 1 in that the molar ratio of NaH2PO4, NaBr, and Bi(NO3)3·5H2O is 2:8:10, resulting in a BiPO4 / BiOBr heterojunction powder of 20 wt%.

[0074] Example 3 differs from Example 1 in that the molar ratio of NaH2PO4, NaBr, and Bi(NO3)3·5H2O is 3:7:10, resulting in a BiPO4 / BiOBr heterojunction powder of 30 wt%.

[0075] Example 4 differs from Example 1 in that the molar ratio of NaH2PO4, NaBr, and Bi(NO3)3·5H2O is 4:6:10, resulting in a BiPO4 / BiOBr heterojunction powder of 40 wt%.

[0076] Example 5 differs from Example 1 in that the molar ratio of NaH2PO4, NaBr, and Bi(NO3)3·5H2O is 5:5:10, resulting in a BiPO4 / BiOBr heterojunction powder of 50 wt%.

[0077] Example 6 differs from Example 1 in that the molar ratio of NaH2PO4, NaBr, and Bi(NO3)3·5H2O is 0:10:10, resulting in a BiPO4 / BiOBr heterojunction powder with a weight of 0 wt%.

[0078] Experimental results:

[0079] The effects of different BiPO4 to BiOBr ratios on the photocurrent of the materials in Examples 1-6 are as follows: Figure 1 As shown, when the BiPO4:BiOBr ratio changes from 0 wt% to 50 wt%, a stronger photocurrent intensity can be obtained when the ratio is between 30-50 wt%, and the peak value is reached when the ratio is 40 wt%. Therefore, the preferred molar ratio of NaBr, NaH2PO4, and Bi(NO3)3·5H2O is (0.5-0.7):(0.5-0.3):1.

[0080] II. Investigation of the concentration of BiPO4 / BiOBr / CdS suspension

[0081] Example 7

[0082] Water-soluble CdS quantum dots were loaded onto a BiPO4 / BiOBr heterojunction using a physical bonding method. First, 2 mg of BiPO4 / BiOBr heterojunction powder and 2 mg of CdS QDs powder were weighed separately, and 4 ml of deionized water was added to dissolve the mixture. The mixture was vortexed for 5 minutes to ensure thorough mixing, resulting in a concentration of 1 mg / mL. -1 The suspension was subjected to ultrasonic treatment for 1 hour to allow the two materials to bond, resulting in a CdS-sensitized BiPO4 / BiOBr composite suspension.

[0083] Example 8 differs from Example 7 in that 4 mg of BiPO4 / BiOBr heterojunction powder and 4 mg of CdSQDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 2 mg / mL. -1 A suspension.

[0084] Example 9 differs from Example 7 in that 6 mg of BiPO4 / BiOBr heterojunction powder and 6 mg of CdSQDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 3 mg / mL. -1 A suspension.

[0085] Example 10 differs from Example 7 in that 8 mg of BiPO4 / BiOBr heterojunction powder and 8 mg of CdSQDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 4 mg / mL. -1 A suspension.

[0086] Example 11 differs from Example 7 in that 10 mg of BiPO4 / BiOBr heterojunction powder and 10 mg of CdS QDs powder were weighed separately, and 4 ml of deionized water was added to dissolve the mixture, preparing a solution with a concentration of 5 mg / mL. -1 A suspension.

[0087] Experimental results:

[0088] The effects of different BiPO4 / BiOBr / CdS suspension concentrations on the photocurrent of the materials in Examples 7-11 are as follows: Figure 3 As shown, the concentration of the BiPO4 / BiOBr / CdS suspension ranges from 1 mg / mL. -1 Change to 5 mg·mL -1 When the concentration is 3 mg·mL -1The photocurrent intensity reaches its peak. This is because when the content of BiPO4 / BiOBr / CdS is too high, the transfer of electrons to the electrode interface is hindered.

[0089] III. Investigation of the mass ratio of BiPO4 / BiOBr and CdS

[0090] Example 12

[0091] Water-soluble CdS quantum dots were loaded onto a BiPO4 / BiOBr heterojunction using a physical bonding method. First, 6 mg of BiPO4 / BiOBr heterojunction powder and 6 mg of CdS QDs powder were weighed separately, and 4 ml of deionized water was added to dissolve the mixture. The mixture was vortexed for 5 minutes to ensure thorough mixing, resulting in a concentration of 3 mg / mL. -1 The suspension has a BiPO4 / BiOBr to CdS mass ratio of 1:1.

[0092] Example 13 differs from Example 12 in that 4 mg of BiPO4 / BiOBr heterojunction powder and 8 mg of CdSQDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 3 mg / mL. -1 The suspension has a BiPO4 / BiOBr to CdS mass ratio of 1:2.

[0093] Example 14 differs from Example 12 in that 3 mg of BiPO4 / BiOBr heterojunction powder and 9 mg of CdSQDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 3 mg / mL. -1 The suspension has a BiPO4 / BiOBr to CdS mass ratio of 1:3.

[0094] Example 15 differs from Example 12 in that 2.4 mg of BiPO4 / BiOBr heterojunction powder and 9.6 mg of CdS QDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 3 mg / mL. -1 The suspension has a BiPO4 / BiOBr to CdS mass ratio of 1:4.

[0095] Example 16 differs from Example 12 in that 2 mg of BiPO4 / BiOBr heterojunction powder and 10 mg of CdS QDs powder were weighed and added to 4 ml of deionized water to dissolve the mixture, preparing a solution with a concentration of 3 mg / mL. -1 The suspension has a BiPO4 / BiOBr to CdS mass ratio of 1:5.

[0096] Experimental results:

[0097] The effect of the BiPO4 / BiOBr and CdS mass ratio on the photocurrent of the materials in Examples 12-16 is as follows: Figure 2 As shown, as the mass ratio of BiPO4 / BiOBr to CdS changes from 1:1 to 1:5, a stronger photocurrent intensity can be obtained when the mass ratio is between 1:3 and 1:5, with the peak value reached at a ratio of 1:3. This is because as the amount of CdS increases, the material is continuously peeled off from the electrode surface, causing a decrease in photocurrent.

[0098] IV. Examination of AA Concentration During Measurement

[0099] Example 17

[0100] Measurement conditions of photocurrent: This study employed a three-electrode system, with the fabricated sensor used as the working electrode; an Ag / AgCl electrode as the reference electrode; and a platinum electrode as the counter electrode. The photocurrent was measured at a concentration of 0.1 mol·L⁻¹. -1 0.1 mol of ascorbic acid (AA) was added to PBS buffer solution as an electron donor to plot the photocurrent. A 250W xenon lamp was used as the excitation source, switched on and off every 20 seconds, with an external voltage of 0.0V.

[0101] Example 18 differs from Example 17 in that 0.2 mol of ascorbic acid (AA) is added to the PBS buffer solution.

[0102] Example 19 differs from Example 17 in that 0.3 mol of ascorbic acid (AA) is added to the PBS buffer solution.

[0103] Example 20 differs from Example 17 in that 0.4 mol of ascorbic acid (AA) is added to the PBS buffer solution.

[0104] Example 21 differs from Example 17 in that ascorbic acid (AA) is not added to the PBS buffer solution.

[0105] Experimental results:

[0106] The effect of AA concentration on the measured photocurrent in Examples 17-21 is as follows: Figure 4 As shown in the figure. It can be seen that as the AA concentration changes from 0 to 0.4 mol·L⁻¹... -1 When the concentration is 0.1 mol·L⁻¹ -1 The photocurrent intensity reaches its peak at this point. The concentration of AA is also an important parameter for antigen capture and specific recognition of signal markers on the electrode surface. However, when the AA concentration is too high, it will hinder electron transfer, thereby causing a decrease in photocurrent.

[0107] V. Investigation of AFP antibody incubation time

[0108] Example 22

[0109] After the PEC composite electrode was successfully prepared, 30 μl of AFP antibody (10 μg·mL⁻¹) was added dropwise. -1 The AFP antibody was then incubated at 37°C for 30 minutes to modify the surface of the BiPO4 / BiOBr / CdS modified electrode.

[0110] Example 23 differs from Example 22 in that the incubation time for the AFP antibody is 40 min.

[0111] Example 24 differs from Example 22 in that the incubation time for the AFP antibody is 50 min.

[0112] Example 25 differs from Example 22 in that the incubation time for the AFP antibody is 60 min.

[0113] Example 26 differs from Example 22 in that the incubation time for the AFP antibody is 70 min.

[0114] Experimental results:

[0115] The incubation time of AFP antibodies also plays a significant role in photocurrent performance measurement. The effects of AFP antibody incubation time on photocurrent performance in Examples 22-26 are as follows: Figure 5 As shown, the photocurrent signal gradually weakens during incubation. When the incubation time for AFP antibody is 70 minutes, the photocurrent signal reaches its minimum, indicating that the immune response is in equilibrium. However, when the incubation time is 60 minutes, the photocurrent decline reaches a plateau. Therefore, to save time, 60 minutes is chosen as the optimal incubation time for AFP antibody.

[0116] VI. Investigation of incubation time for different concentrations of AFP

[0117] Example 27

[0118] 30 μl of BSA (1 wt%) was added dropwise to the surface of the modified electrode to block non-specific binding sites. 0.01 mol·L⁻¹ -1 The modified electrode was thoroughly cleaned with PBS buffer solution. Then, 30 μl of AFP antigen at different concentrations was evenly dropped onto the surface of the modified electrode and incubated at 37 °C for 30 min, thereby modifying the surface of the BiPO4 / BiOBr / CdS modified electrode with different concentrations of AFP antigen.

[0119] Example 28 differs from Example 27 in that the incubation time for the AFP antigen is 40 minutes.

[0120] Example 29 differs from Example 27 in that the incubation time for the AFP antigen is 50 min.

[0121] Example 30 differs from Example 27 in that the incubation time for the AFP antigen is 60 min.

[0122] Example 31 differs from Example 27 in that the incubation time for the AFP antigen is 70 min.

[0123] Experimental results:

[0124] The incubation time of AFP antigen also plays a significant role in photocurrent performance measurement. The effect of AFP antigen incubation time on photocurrent performance in Examples 27-31 is as follows: Figure 6 As shown, the photocurrent signal gradually weakens during incubation. When the incubation time for AFP antigen is 60 minutes, the photocurrent signal reaches its minimum value, indicating that the immune response is in equilibrium. Therefore, 60 minutes is the optimal incubation time for AFP antibodies.

[0125] VII. Detection of AFP Concentration in Different Samples Using Photocurrent Intensity

[0126] The following experiments were conducted using the preferred results from Examples 1-31 above.

[0127] Example 32

[0128] Step 1: Preparation of BiPO4 / BiOBr composite material;

[0129] BiPO4 / BiOBr composites were synthesized via a one-step hydrothermal method. Under stirring conditions, 0.1235 g NaBr, 0.096 g NaH2PO4, and 0.97 g Bi(NO3)3·5H2O (molar ratio NaBr:NaH2PO4:Bi(NO3)3·5H2O = 0.6:0.4:1) were simultaneously mixed in 60 ml of methanol and stirred vigorously for 15 min. The mixture was then sonicated for 15 min, stirred again at room temperature for 30 min, and then transferred to a polytetrafluoroethylene-lined reactor and heated at 180 °C for 24 h. After the reactor cooled naturally, the precipitate was collected by centrifugation, and the product was washed three times with anhydrous ethanol and ultrapure water. Finally, the precipitate was dried in a forced-air drying oven at 60 °C for 12 h to obtain BiPO4 / BiOBr heterojunction powder for subsequent experiments.

[0130] Step 2: Preparation of water-soluble CdS quantum dots;

[0131] 0.2284 g of CdCl2 and 100 ml of deionized water were placed in a 250 ml three-necked round-bottom flask to prepare 0.01 mol·L⁻¹ CdCl₂. - 1 CdCl2 solution. After adding 500 μl of mercaptopropionic acid, the solution was degassed with nitrogen for 30 min, and then treated with 1.0 mol·L⁻¹.-1 Adjust the pH to 11 with NaOH solution. Finally, add 10 ml of 0.1 mol·L⁻¹ NaOH solution to the mixed solution. -1 The Na₂S solution was heated under reflux at 110°C for 4 hours. After the solution cooled to room temperature, 25 ml of isopropanol was added for precipitation, and the solid precipitate was obtained by centrifugation. The precipitate was washed three times with anhydrous ethanol and dried in a vacuum drying oven at 60°C for 12 hours to obtain CdS QDs powder for subsequent experiments.

[0132] Step 3: Preparation of BiPO4 / BiOBr / CdS composite material;

[0133] Water-soluble CdS quantum dots were loaded onto a BiPO4 / BiOBr heterojunction using a physical bonding method. First, 3 mg of BiPO4 / BiOBr heterojunction powder and 9 mg of CdS QDs powder were weighed out. Next, 4 mL of deionized water was added to dissolve the mixture, and the mixture was vortexed for 5 min to ensure thorough mixing. After ultrasonic treatment for 1 h to allow the two materials to bond, a CdS-sensitized BiPO4 / BiOBr composite suspension (concentration 3 mg / mL) was obtained. -1 Store it in a refrigerator at 4°C away from light for later use.

[0134] Step 4: Preparation of the PEC immunosensor;

[0135] First, the CdS-sensitized BiPO4 / BiOBr composite suspension prepared in the previous step was uniformly added dropwise to FTO, and dried in a vacuum drying oven at 60°C for 12 hours. At this point, the composite electrode was successfully prepared. Then, 30 μl of AFP antibody (10 μg / mL) was added dropwise. -1 The AFP antibody was incubated at 37°C for 60 min to modify the surface of the BiPO4 / BiOBr / CdS modified electrode. Next, 30 μl of BSA (1 wt%) was added dropwise to the surface of the modified electrode to block non-specific binding sites. After 90 min, 30 μl of a 0.001 ng / mL solution was added. -1 The AFP antigen analyte was uniformly dropped onto the surface of the modified electrode and incubated for 60 min to induce specific immune binding. The sensor was prepared at room temperature, and the PEC immunosensor was stored at 4°C until testing. To remove excess free protein, the modified electrode was thoroughly washed with PBS buffer after each modification.

[0136] Example 33 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 0.005 ng·mL⁻¹. 1 .

[0137] Example 34 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 0.01 ng / mL.-1 .

[0138] Example 35 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 0.05 ng / mL. -1 .

[0139] Example 36 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 0.1 ng / mL. -1 .

[0140] Example 37 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 0.5 ng / mL. -1 .

[0141] Example 38 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 1 ng / ml. -1 .

[0142] Example 39 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 5 ng / mL. -1 .

[0143] Example 40 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 10 ng / mL. -1 .

[0144] Example 41 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 50 ng / mL. -1 .

[0145] Example 42 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 100 ng / mL. -1 .

[0146] Example 43 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 500 ng / mL. -1 .

[0147] Example 44 differs from Example 32 in that the concentration of AFP antigen added to the modified electrode is 1000 ng·mL⁻¹. 1 .

[0148] The application of the prepared PEC immunosensor in the detection of alpha-fetoprotein includes the following steps:

[0149] (1) A three-electrode system was used: the prepared PEC immunosensor was used as the working electrode; the Ag / AgCl electrode was used as the reference electrode; and the platinum electrode was used as the counter electrode. The supernatant of the human serum sample was taken as the initial sample after centrifugation three times and stored in a refrigerator at 4°C.

[0150] (2) At a concentration of 0.1 mol·L -1 Ascorbic acid (AA) was added to the PBS buffer solution as an electron donor.

[0151] (3) The excitation light source is a 250W xenon lamp, which is switched on and off every 20 seconds, with an external voltage of 0.0V.

[0152] (4) The photocurrent curves of the working electrode under illumination and non-illumination conditions were collected using an electrochemical workstation. The xenon lamp was switched on and off once every 20 seconds as one cycle, and the difference between the highest and lowest points on the vertical axis was taken as the photocurrent intensity.

[0153] (5) Plot the standard curve about AFP concentration: Plot the standard curve about AFP concentration with the photocurrent intensity of the sensor as the vertical axis and the AFP concentration as the horizontal axis to obtain the standard curve graph.

[0154] (6) Detection of AFP concentration in unknown solution: The unknown solution is incubated on the prepared PEC immunosensor, the photocurrent intensity of the sensor is detected, and the concentration of AFP in the unknown solution can be obtained by combining the standard curve.

[0155] Experimental results:

[0156] The correction equation was obtained by linearly fitting the logarithm of the antigen concentration to the photocurrent intensity. The fitting result is as follows: Figure 7 As shown, the obtained regression equation is I = -0.2775lgC AFP (ng·mL -1 )+1.919(R 2 =0.999). Because the immune complex formed on the electrode surface acts as an inert barrier layer hindering electron transfer, the photocurrent signal is limited to 0.001-1000 ng / mL. -1 Within the range, the concentration of AFP decreased with increasing concentration, showing a good linear relationship; the LOD (limit of detection) was calculated to be 0.82 pg·mL. -1 It has good sensitivity and accuracy.

[0157] VII. Specificity of this method determined by photocurrent intensity.

[0158] Specificity is crucial for most immunosensors because nonspecific binding can mislead detection results. The PEC immunosensor prepared in Example 32 was used to perform a specific recognition experiment on biomarkers that may be present in serum. Unlike Example 32, the analyte incubated on the PEC immunosensor was at a concentration of 10 μg / mL. -1 Interfering substances and concentrations of 100 ng·mL -1 Add 10 μg·mL of AFP -1The interfering agents were: a: blank; b: carcinoembryonic antigen (CEA); c: human hemoglobin; d: alkaline phosphatase; e: human serum albumin; f: adenosine triphosphate (ATP); g: cytochrome C; h: AFP; i: AFP + CEA; j: AFP + human hemoglobin; k: AFP + alkaline phosphatase; l: AFP + human serum albumin; m: AFP + ATP; n: AFP + cytochrome C. After incubation, the modified electrodes were thoroughly washed with PBS buffer, and the photocurrent intensity was measured.

[0159] Test results as follows Figure 8 As shown, in the absence of AFPs (a, b, c, d, e, f, g), incubation with the interfering substances did not cause significant changes in the photocurrent on the electrodes. However, in the presence of AFPs (h, i, j, k, l, m, n), no obvious photocurrent fluctuations were observed after incubation with the aforementioned interfering substances on the electrodes. Therefore, the presence of the interfering proteins does not significantly affect the photocurrent response of this PEC immunosensor, indicating that the PEC immunosensor has excellent selectivity and specificity for AFPs.

[0160] VIII. Repeatability and Stability of the Method Based on Photocurrent Intensity Measurement

[0161] Repeatability and stability are also important characteristics of PEC immunosensors, and are of great significance for the development and application of biosensors. Five PEC immunosensors prepared in Example 32 were used, with a detection concentration of 100 ng / mL. -1 The AFP performed repeatability tests on the constructed sensor.

[0162] The results are as follows Figure 9 As shown, the photocurrent intensities of the five PEC immunosensors measured were similar, and the calculated relative standard deviation (RSD) was 2.13%, providing evidence of good repeatability of the sensors.

[0163] The PEC immunosensor prepared in Example 32 was used to detect a concentration of 100 ng / mL. -1 The stability of the constructed sensor was tested using an AFP. After multiple on / off illumination cycles within 400 seconds, the change in photocurrent intensity was observed.

[0164] The results are as follows Figure 10 As shown, after multiple on / off irradiation cycles within 400 seconds, the photocurrent remains unchanged, indicating that the sensor has good stability.

[0165] A series of results indicate that the PEC immunosensor has good repeatability and stability.

[0166] IX. Determination of AFP concentration in serum samples using photocurrent intensity

[0167] The feasibility and applicability of the PEC immunosensor prepared by this method in determining AFP levels in human serum were investigated using the method described in Example 32. Unlike Example 32, the analyte incubated on the PEC immunosensor was serum samples containing different concentrations of AFP. Fresh human serum was centrifuged at 12,000 rpm for 20 min, and the supernatant was diluted 50-fold with deionized water to obtain pure serum samples. Relatively low, medium, and high concentrations of AFP solutions were sequentially introduced into the diluted serum samples, and the spiked solutions were analyzed using the same procedure and conditions as in Example 32. The experimental results are shown in Table 1. The average recovery rate of AFP was 97.4%–104.9%, and the relative standard deviation did not exceed 2.4%. The results indicate that the PEC immunosensor prepared by this method is feasible for determining AFP in serum, has good detection sensitivity, and has good application value and prospects in the fields of biology and medicine.

[0168] Table 1. AFP concentration in serum samples determined by photocurrent intensity.

[0169]

[0170]

[0171] "-" indicates that it was not detected.

[0172] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. The present invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be within the protection scope of the present invention.

Claims

1. A method for preparing a ternary heterojunction composite photoelectrochemical immunosensor, characterized in that, The heterojunction is a BiPO4 / BiOBr / CdS ternary heterojunction composite material, comprising the following steps: Step 1: preparation of BiPO4 / BiOBr composite material; Under stirring conditions, NaBr, NaH2PO4 and Bi(NO3)3·5H2O are added to the solvent for mixing, and the mixture is stirred vigorously to obtain a mixed solution, which is then ultrasonically treated and continuously stirred at room temperature, and then transferred to a polytetrafluoroethylene-lined reaction kettle for heating reaction, and after the reaction is completed, the mixture is naturally cooled, the precipitate is collected by centrifugation, and the precipitate is washed with anhydrous ethanol and ultrapure water, respectively, and then dried to obtain a BiPO4 / BiOBr heterojunction powder; Step 2: preparation of water-soluble CdS QDs; CdCl2 is added to deionized water to prepare a CdCl2 solution, and after adding mercaptopropionic acid, the solution is degassed with nitrogen, and then the pH value is adjusted to 9-12 with a NaOH solution, and a Na2S solution is added, and the mixture is heated to reflux for 2-6 h, and then the solution is cooled to room temperature, and isopropanol is added for sedimentation treatment, and the solid precipitate is obtained by centrifugation, and the precipitate is washed with anhydrous ethanol and dried to obtain a CdS QDs powder; Step 3: preparation of a BiPO4 / BiOBr / CdS ternary heterojunction composite material; BiPO4 / BiOBr heterojunction powder and CdS QDs powder are weighed separately, deionized water is added, and the mixture is vortexed to obtain a mixed solution, which is ultrasonically treated for 30-90 min to combine the two materials, and a CdS-sensitized BiPO4 / BiOBr composite suspension is obtained, which is stored at 4°C in the dark for later use; Step 4: preparation of a PEC immunosensor; The FTO glass electrode is cleaned, and the CdS-sensitized BiPO4 / BiOBr / CdS suspension is uniformly dropped onto the glass electrode, which is then dried, and AFP antibody, BSA and different concentrations of AFP antigen are added to the dried glass electrode.

2. The method for preparing a ternary heterojunction composite photoelectrochemical immunosensor according to claim 1, characterized in that, In step 1, the molar ratio of NaBr, NaH2PO4 and Bi(NO3)3·5H2O is (0.5-0.9):(0.5-0.1):

1.

3. The method for preparing a ternary heterojunction composite photoelectrochemical immunosensor according to claim 1, characterized in that, In step 1, the mixture is heated in a polytetrafluoroethylene-lined reaction kettle at 160-220°C for 12-24 h, and the obtained precipitate is dried at 40-80°C for 12-24 h to obtain a BiPO4 / BiOBr heterojunction powder with a mass percentage of 10-50 wt%.

4. The method for preparing a ternary heterojunction composite photoelectrochemical immunosensor according to claim 1, characterized in that, In step 2, the molar ratio of CdCl2 to Na2S in the mixed solution is 1:(0.5-1), and the material ratio of CdCl2 to mercaptopropionic acid is 100-500 μl of mercaptopropionic acid per 0.001 mol of CdCl2, and the concentration of the Na2S solution is 0.1 mol / L. -1 .

5. The method for preparing a ternary heterojunction composite photoelectrochemical immunosensor according to claim 1, characterized in that, In step 3, the mass ratio of BiPO4 / BiOBr heterojunction powder and CdS QDs powder is 1:1-1:

5.

6. The method for preparing a ternary heterojunction composite photoelectrochemical immunosensor according to claim 1, characterized in that, In step 3, the concentration of the obtained CdS-sensitized BiPO4 / BiOBr / CdS composite suspension is 1.0-5.0 mg·mL -1 .

7. The method according to claim 1, wherein the method is characterized by, The FTO glass electrode used in step 4 has a size of 0.6 x 2.0 cm 2 The cleaning process uses acetone, a mixed solution of 1.0 mol·L -1 -1 NaOH and 50% ethanol, and ultrapure water, respectively, and the cleaned FTO glass electrode is dried at 40-80 ℃ for 2-6 h; 10-50 μl of AFP antibody with a concentration of 5-20 μg·mL -1 -1 is added dropwise to the surface of the BiPO4 / BiOBr / CdS modified electrode; then, 10-50 μl of BSA solution with a concentration of 0.5-2 wt% is added dropwise to the surface of the modified electrode; finally, 10-50 μl of AFP antigen with a concentration of 0.001-1000 ng·mL -1 -1 is uniformly dropped on the surface of the modified electrode.

8. A ternary heterojunction composite photoelectrochemical immunosensor, characterized in that, Obtained by the preparation method of any one of claims 1-7.

9. Use of the ternary heterojunction composite material photoelectrochemical immunosensor of claim 8 for detecting alpha-fetoprotein, specifically comprising the following steps: (1) Three-electrode system was adopted: the prepared ternary heterojunction composite material photoelectrochemical immunosensor was used as the working electrode; Ag / AgCl electrode was used as the reference electrode; platinum electrode was used as the counter electrode; human serum sample was centrifuged for 3 times, and the supernatant was taken as the initial sample, which was stored in a refrigerator at 4 DEG C; (2) Ascorbic acid was added as an electron donor in a PBS buffer solution with a concentration of 0.1 mol·L -1 -1. (3) The excitation light source was a 250 W xenon lamp, which was turned on and off every 20 s, and an external voltage of 0.0 V was applied; (4) The electrochemical workstation was used to collect the photocurrent curves of the working electrode under light and non-light conditions, and the difference between the highest point and the lowest point of the vertical coordinate was taken as the photocurrent intensity, with the xenon lamp turned on and off every 20 s as a cycle; (5) The standard curve of alpha-fetoprotein concentration was drawn: the photocurrent intensity of the sensor was taken as the vertical coordinate, and the alpha-fetoprotein concentration was taken as the horizontal coordinate, to draw the standard curve of alpha-fetoprotein concentration, and obtain the standard curve graph; (6) The alpha-fetoprotein concentration in the unknown solution was detected: the unknown solution was incubated on the prepared ternary heterojunction composite material photoelectrochemical immunosensor, the photocurrent intensity of the sensor was detected, and the alpha-fetoprotein concentration in the unknown solution could be obtained combined with the standard curve graph.

10. Use of a ternary heterojunction composite photoelectrochemical immunosensor according to claim 9 for the detection of alpha-fetoprotein, characterized in that, The linear range of the ternary heterojunction composite photoelectrochemical immunosensor for alpha-fetoprotein is 0.001-1000 ng·mL -1 , and the detection limit is 0.82 pg·mL -1 .

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