CN110501401B - Preparation method of photoelectrochemical immunosensor based on bismuth molybdate/zinc-doped cadmium sulfide/gold - Google Patents
Preparation method of photoelectrochemical immunosensor based on bismuth molybdate/zinc-doped cadmium sulfide/gold Download PDFInfo
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Abstract
The invention relates toA preparation method of a photoelectrochemical immunosensor based on bismuth molybdate/zinc doped cadmium sulfide/gold. According to the invention, zinc-doped cadmium sulfide sensitized bismuth molybdate nanosheets are used as photosensitive substrate materials, bismuth molybdate is simple to prepare, and has a good photocatalytic effect, and good photoelectric response is obtained after the zinc ion doped cadmium sulfide is sensitized. The gold nanoparticles are adopted to further modify the sensitized substrate photosensitive material, so that on one hand, the band gap defect of cadmium sulfide is improved after zinc doping, the surface plasma effect of gold is more remarkable, the vibration of electrons on the surface of the photosensitive material is promoted, and the electron transfer is accelerated to obtain more stable and excellent photocurrent; on the other hand, Au-NH is easily formed between gold and biomolecules2The bond provides a foundation for the connection of inorganic semiconductor nano materials and biomolecules, and improves the stability and sensitivity of the sensor. The detection limit of the sensor reaches 0.03 pg/mL.
Description
Technical Field
The invention relates to a preparation method of a photoelectrochemical immunosensor based on bismuth molybdate/zinc doped cadmium sulfide/gold. Specifically, zinc-doped cadmium sulfide sensitized bismuth molybdate nano-sheets are used as a substrate photosensitive material to provide basic photoelectric response, noble metal gold is used for further modifying sensitization, on one hand, the photocurrent is improved due to the surface plasmon resonance effect, and on the other hand, the gold and biomolecules form stable Au-NH2And a foundation is provided for the construction of a subsequent biological immune sensing platform. A standard-free photoelectrochemical sensor for detecting procalcitonin is prepared, and belongs to the technical field of novel functional materials and biosensing detection.
Background
Procalcitonin (PCT) is a protein whose levels in plasma are elevated when severe bacterial, fungal and parasitic infections and sepsis and multi-organ failure, bacterial endotoxins play a crucial role in the induction process, and localized limited bacterial infections, mild infections and chronic inflammation do not lead to an elevation, nor does PCT rise in autoimmune, allergic and viral infections, and thus PCT is a specific indicator of severe bacterial and fungal inflammation and also a reliable indicator of sepsis and multi-organ failure associated with inflammatory activity, reflecting the activity of systemic inflammatory reactions. In severe bacterial infectious diseases such as sepsis and MODS, the degree of PCT elevation is a reflection of inflammatory activity. PCT is not only an acute indicator for differential diagnosis, but also a parameter for monitoring inflammatory activity. Its advantage over other inflammatory markers is that severe infections cause significant increases in PCT concentrations (> 10 ug/L), whereas less severe infections or less severe sepsis cause only moderate increases in PCT. Further, PCT can also be used as a post-cure monitoring indicator of severe inflammation. PCT returned to the normal reference range within a few days if the inflammatory stimulus was no longer present. Therefore, it is necessary to establish an analysis method for rapidly and accurately detecting procalcitonin. There are many existing methods for detecting procalcitonin antigen, such as the detection by time-resolved fluorescence immunochromatography (Huang De Zhi, Yihao, Liufei, etc.. the establishment and performance evaluation of procalcitonin time-resolved fluorescence immunochromatography detection method [ J ]. third Jun Med. academic newspaper, 2019, 41(06): 581-); enzyme-linked immunosorbent assay (enzyme-linked immunosorbent assay for procalcitonin, CN 201610401787.1); chemiluminescence immune methods (procalcitonin chemiluminescence detection reagent and detection method based on nano-antibody, CN201810235448. X) and the like, but enzyme-linked immunoassay is complicated in operation and expensive in price; the controllability of fluorescence analysis is poor, and the toxicity is high; long detection time of electrochemical luminescence analysis, and the like. The invention designs a novel standard-free photoelectrochemical sensor which has the advantages of high analysis speed, simple operation, good stability and low detection limit, and the detection limit of the standard-free photoelectrochemical sensor designed by the invention on procalcitonin antigen reaches 0.03 pg/mL.
Bismuth molybdate is a typical semiconductor photocatalysis nano material, has the advantages of simple preparation, low cost, no toxicity and no pollution, has good absorption and utilization efficiency on visible light, has certain photostability, can generate photoproduction charges under the irradiation of the visible light so as to form photocurrent, but is easy to generate photoproduction electron holes due to narrow band gapIn combination, the photoelectric conversion efficiency is not high. Cadmium sulfide is used as a good sensitizer, so that the photoelectric defect of bismuth molybdate can be well improved, and after the sensitization by cadmium sulfide, the photoelectric response of bismuth molybdate is greatly improved, so that a foundation is provided for subsequent photoelectric tests. In addition, after zinc ion doping, a middle band gap structure is formed in cadmium sulfide, the recombination rate of photo-generated electron holes of the cadmium sulfide is reduced, and the stability of the sensor is improved. Secondly, gold nanoparticles are adopted for further modification and sensitization, so that on one hand, the gold has a good surface plasmon resonance effect, and can accelerate the vibration of electrons on the surface of the substrate material and promote the rapid transfer of the electrons; on the other hand, Au-NH can be easily formed between gold and biomolecules2The bond realizes the purpose of connecting the inorganic semiconductor material and the biological molecules without adding any cross-linking agent, and provides a foundation and guarantee for the construction of the immunosensing platform.
A photoelectrochemical sensor is a type of detection device that determines the concentration of an analyte based on the photoelectric conversion characteristics of a substance. The photoelectrochemical detection method has the characteristics of simple equipment, high sensitivity and easiness in miniaturization, has been developed into an analysis method with great application potential, and has wide application prospects in the fields of food, environment, medicine and the like. The application of the bismuth molybdate material in the aspect of the photoelectrochemical sensor is not reported. The invention successfully constructs a standard-free photoelectrochemical sensor for detecting procalcitonin under visible light based on bismuth molybdate/zinc-doped cadmium sulfide/gold nanoparticle materials. The sensor takes zinc-doped cadmium sulfide sensitized flaky bismuth molybdate as a substrate photosensitive material, is further modified by gold nanoparticles, and realizes sensitive detection of procalcitonin according to specific immune recognition of antigen and antibody. The photoelectrochemical sensor prepared by the invention has the advantages of low cost, high sensitivity, good specificity, quick detection, easy preparation and the like, realizes quick and high-sensitivity detection of procalcitonin in a visible light region, and effectively overcomes the defects of the existing procalcitonin detection method.
Disclosure of Invention
One of the purposes of the invention is to adopt a flaky bismuth molybdate nano semiconductor material as a photosensitive substrate, wherein the material has a large specific surface area and shows satisfactory photoelectric response under visible light.
The other purpose of the invention is to adopt zinc-doped cadmium sulfide nanoparticles as a sensitizer to sensitize flaky bismuth molybdate materials, the flaky structure of the bismuth molybdate can be loaded with a large amount of sensitizer, and the band gap defect of cadmium sulfide is improved after zinc ions are doped, so that the cadmium sulfide and the bismuth molybdate are in band gap matching, the electron transfer is quicker, excellent photocurrent response can be obtained, and the sensitivity of the sensor is improved.
The third purpose of the invention is to adopt noble metal gold nanoparticles to further sensitize and modify a substrate photosensitive material, so that on one hand, the gold has a good surface plasma resonance effect, so that the electronic vibration on the surface of the photosensitive material is more severe, and the transfer rate of electrons is improved; on the other hand, the noble metal gold and the biological molecules can form firm Au-NH2The bond can realize the combination of the inorganic semiconductor material and the biological molecule without adding a cross-linking agent, and provides a foundation for the construction of an immunosensing platform.
The fourth purpose of the invention is to use zinc-doped cadmium sulfide sensitized bismuth molybdate nano-sheets as a substrate, and further modify and sensitize gold nano-particles to prepare the photoelectrochemical sensor with high sensitivity, good stability and high detection speed, thereby realizing the purpose of sensitively detecting procalcitonin under the condition of visible light.
The technical scheme of the invention is as follows:
1. a preparation method of a photoelectrochemical immunosensor based on bismuth molybdate/zinc-doped cadmium sulfide/gold is characterized by comprising the following steps:
(1) preparation of bismuth molybdate nanosheet material
Dissolving 1.0-2.0 g of pentahydrate bismuth nitrate and 0.4-0.6 g of sodium molybdate dihydrate into 100-150 mL of ultrapure water under stirring, stirring the solution at room temperature for 1-3 h, transferring the uniformly stirred pale yellow solution into a high-pressure reaction kettle, reacting at 160-200 ℃ for 20-24 h, naturally cooling to room temperature after the reaction is finished, washing the product with ultrapure water and absolute ethyl alcohol for 3 times respectively, and then vacuum-drying at 30-50 ℃ for 8-12 h to prepare the flaky bismuth molybdate nano material;
(2) preparation of zinc-doped cadmium sulfide nanoparticles
Taking zinc acetate dihydrate and cadmium acetate dihydrate according to different proportions, co-dissolving the zinc acetate dihydrate and the cadmium acetate dihydrate in 20-50 mL of ultrapure water to ensure that the total molar mass of zinc ions and cadmium ions is 0.01 mol, and uniformly stirring the solution at room temperature to obtain a solution A; dissolving 1.0-3.0 g of sodium sulfide nonahydrate in 20-50 mL of ultrapure water, and uniformly stirring to obtain a solution B; mixing the solution A and the solution B, stirring for 6-12 h at room temperature, then pouring the mixed aqueous solution into a high-pressure reaction kettle, reacting for 14-18 h at 150-200 ℃, naturally cooling to room temperature after the reaction is finished, washing the product for 3 times by using ultrapure water and absolute ethyl alcohol respectively, and then carrying out vacuum drying overnight at 30-50 ℃ to obtain cadmium sulfide nano-particle materials with different zinc doping ratios;
(3) preparation of gold nanoparticles
Boiling 30-50 mL of chloroauric acid aqueous solution with the mass fraction of 0.01-0.05% at 120 ℃ for 5-10 min, then adding 2.0-5.0 mL of trisodium citrate aqueous solution with the mass fraction of 1-3% into the boiled solution, continuously stirring the solution under the boiling condition, changing the color of the solution from slight blue to bright red, and continuously stirring to obtain a purple solution, thus obtaining gold nanoparticles;
(4) preparation of photoelectrochemical sensor
1) Ultrasonically cleaning conductive glass by using a detergent, acetone, ethanol and ultrapure water in sequence, and drying by using nitrogen;
2) dripping 6 muL and 2-6 mg/mL bismuth molybdate water solution onto a conductive surface of the ITO conductive glass, and naturally airing at room temperature;
3) continuously dropwise adding zinc-doped cadmium sulfide aqueous solution of 6 muL and 2-6 mg/mL on the surface of the modified electrode, and naturally airing;
4) continuously dropwise adding 6 mu L of gold nanoparticle aqueous solution on the surface of the modified electrode, and naturally airing at room temperature;
5) dropwise adding 4 muL of procalcitonin antibody of 5-20 mug/mL, washing with ultrapure water, and naturally airing at room temperature to a wet film state;
6) dropwise adding a bovine serum albumin solution with the mass fraction of 0.1% -0.3% and prepared by using a PBS buffer solution with the pH of 7.0 to the surface of the modified electrode, washing the surface of the electrode by using ultrapure water, and airing in a refrigerator at 4 ℃;
7) dropwise adding 6 mu L of procalcitonin antigen, using 0.1 pg/mL-1000 ng/mL of PBS buffer solution for preparation, washing the surface of the electrode with ultrapure water, and naturally airing in a refrigerator at 4 ℃ to prepare the photoelectric chemical sensor for detecting the procalcitonin antigen.
2. The detection method of the photoelectrochemical sensor comprises the following steps:
(1) testing by using an electrochemical workstation and a three-electrode system, taking a saturated calomel electrode as a reference electrode, a platinum wire electrode as an auxiliary electrode, and taking the prepared ITO modified sensor as a working electrode, wherein the testing is carried out in 10 mL of PBS (phosphate buffer solution) containing 0.01-0.5 mol/L ascorbic acid and having the pH value of 5.0-8.0;
(2) detecting the procalcitonin antigen by a time-current method, setting the voltage to be-0.1V, the running time to be 120s, and the wavelength of a light source to be 400-450 nm;
(3) after the electrodes are placed, turning on the lamp every 20s for continuously irradiating for 20s, recording the photocurrent, and drawing a working curve;
(4) and replacing the procalcitonin antigen sample solution to be detected with the procalcitonin antigen standard solution for detection.
The linear range of the sensor for detecting procalcitonin is 0.1 pg/mL-1000 ng/mL, and the detection limit is 0.03 pg/mL.
The chemicals required for material synthesis were all purchased from local reagent stores and were not reprocessed.
Advantageous results of the invention
(1) The invention successfully synthesizes the flaky bismuth molybdate semiconductor nano material with certain photoelectric properties, and the material has the advantages of low cost, no toxicity and large specific surface area; the bismuth molybdate is sensitized by utilizing the good sensitization of cadmium sulfide, so that the excellent photoelectric performance is obtained, and the problem of low photoelectric conversion efficiency of pure bismuth molybdate and pure cadmium sulfide is solved.
(2) According to the invention, zinc ion doped cadmium sulfide is adopted as a sensitizer, the cadmium sulfide has good sensitization, but the band gap of the cadmium sulfide is narrow, the photo-generated electron hole recombination is rapid, and after the zinc ion doping, the band gap defect of the cadmium sulfide is improved, the band gap width of the cadmium sulfide is increased, and the sensitization effect is improved.
(3) The gold nanoparticles are adopted to further modify the sensitized substrate photosensitive material, so that on one hand, the gold has a good surface plasmon resonance effect, can promote vibration of electrons on the surface of the photosensitive material, and promotes rapid transfer of the electrons; on the other hand, stable Au-NH can be formed between the noble metal gold and the biomolecules2The bond combines the inorganic semiconductor material with the biological molecule without adopting any external cross-linking agent, and provides a foundation for the construction of the immunosensing platform.
(4) The photoelectrochemical sensor prepared by the invention is used for detecting procalcitonin, has short response time, wide linear range, low detection limit, good stability and reproducibility, and can realize simple, quick, high-sensitivity and specific detection. The linear range of the procalcitonin detection method is 0.1 pg/mL-1000 ng/mL, and the detection limit is 0.03 pg/mL.
Detailed description of the preferred embodiments
EXAMPLE 1 preparation of photoelectrochemical sensor
(1) Preparation of bismuth molybdate nanosheet material
Dissolving 1.0g of pentahydrate bismuth nitrate and 0.4 g of sodium molybdate dihydrate into 100 mL of ultrapure water under stirring, stirring the solution at room temperature for 1 h, transferring the obtained light yellow solution after uniform stirring into a high-pressure reaction kettle, reacting at 160 ℃ for 20 h, naturally cooling to room temperature after the reaction is finished, washing the product with ultrapure water and absolute ethyl alcohol for 3 times respectively, and then drying in vacuum at 30 ℃ for 8 h to prepare the flaky bismuth molybdate nano material;
(2) preparation of zinc-doped cadmium sulfide nanoparticles
Proportionally taking zinc acetate dihydrate and cadmium acetate dihydrate to dissolve in 20 mL of ultrapure water together, so that the molar mass of zinc ions and cadmium ions is 0.01 mol in total, and uniformly stirring the solution at room temperature to obtain a solution A; dissolving 1.0g of sodium sulfide nonahydrate in 20 mL of ultrapure water, and uniformly stirring to obtain a solution B; mixing the solution A and the solution B, stirring for 6 hours at room temperature, then pouring the mixed aqueous solution into a high-pressure reaction kettle, reacting for 14 hours at 150 ℃, naturally cooling to room temperature after the reaction is finished, washing the product for 3 times by using ultrapure water and absolute ethyl alcohol respectively, and then carrying out vacuum drying at 30 ℃ overnight to obtain the zinc-doped cadmium sulfide nano-particle material;
(3) preparation of gold nanoparticles
Boiling 30 mL of chloroauric acid aqueous solution with the mass fraction of 0.01% at 120 ℃ for 5 min, then adding 2.0 mL of trisodium citrate aqueous solution with the mass fraction of 1% into the boiled solution, continuously stirring the solution under the boiling condition, changing the color of the solution from slight blue to bright red, and continuously stirring to obtain purple solution to prepare gold nanoparticles;
(4) preparation of photoelectrochemical sensor
1) Ultrasonically cleaning conductive glass by using a detergent, acetone, ethanol and ultrapure water in sequence, and drying by using nitrogen;
2) dripping 6 mu L of 2 mg/mL bismuth molybdate aqueous solution onto a conductive surface of the ITO conductive glass, and naturally airing at room temperature;
3) continuously dropwise adding 6 muL and 2 mg/mL zinc-doped cadmium sulfide aqueous solution on the surface of the modified electrode, and naturally airing;
4) continuously dropwise adding 6 mu L of gold nanoparticle aqueous solution on the surface of the modified electrode, and naturally airing at room temperature;
5) dropwise adding 4 muL of procalcitonin antibody of 5 mug/mL, washing with ultrapure water, and naturally airing at room temperature to a wet film state;
6) dropwise adding a bovine serum albumin solution with the mass fraction of 0.1% and prepared by using a PBS buffer solution with the pH of 7.0 to the surface of the modified electrode, washing the surface of the electrode by using ultrapure water, and airing in a refrigerator at 4 ℃;
7) dropwise adding 6 mu L of procalcitonin antigen, using 0.1 pg/mL-1000 ng/mL of PBS buffer solution for preparation, washing the surface of the electrode with ultrapure water, and naturally airing in a refrigerator at 4 ℃ to prepare the photoelectric chemical sensor for detecting the procalcitonin antigen.
EXAMPLE 2 preparation of photoelectrochemical sensor
(1) Preparation of bismuth molybdate nanosheet material
Dissolving 1.5 g of pentahydrate bismuth nitrate and 0.5 g of sodium molybdate dihydrate in 120 mL of ultrapure water under stirring, stirring the solution at room temperature for 2 h, transferring the uniformly stirred pale yellow solution into a high-pressure reaction kettle, reacting at 180 ℃ for 22 h, naturally cooling to room temperature after the reaction is finished, washing the product with ultrapure water and absolute ethyl alcohol for 3 times respectively, and then vacuum-drying at 40 ℃ for 10 h to obtain the flaky bismuth molybdate nano material;
(2) preparation of zinc-doped cadmium sulfide nanoparticles
Proportionally dissolving zinc acetate dihydrate and cadmium acetate dihydrate in 40 mL of ultrapure water to ensure that the molar mass of zinc ions and cadmium ions is 0.01 mol in total, and uniformly stirring the solution at room temperature to obtain a solution A; dissolving 2.0g of sodium sulfide nonahydrate in 30 mL of ultrapure water, and uniformly stirring to obtain a solution B; mixing the solution A and the solution B, stirring for 8 hours at room temperature, then pouring the mixed aqueous solution into a high-pressure reaction kettle, reacting for 16 hours at 180 ℃, naturally cooling to room temperature after the reaction is finished, washing the product for 3 times by using ultrapure water and absolute ethyl alcohol respectively, and then carrying out vacuum drying at 40 ℃ overnight to obtain the zinc-doped cadmium sulfide nano-particle material;
(3) preparation of gold Nanocells
Boiling 40 mL of chloroauric acid aqueous solution with the mass fraction of 0.04% at 120 ℃ for 6 min, then adding 4 mL of trisodium citrate aqueous solution with the mass fraction of 2% into the boiled solution, continuously stirring the solution under the boiling condition, changing the color of the solution from slight blue to bright red, and continuously stirring to obtain purple solution, thus obtaining gold nanoparticles;
(4) preparation of photoelectrochemical sensor
1) Ultrasonically cleaning conductive glass by using a detergent, acetone, ethanol and ultrapure water in sequence, and drying by using nitrogen;
2) dripping 6 mu L of 4 mg/mL of bismuth molybdate aqueous solution onto a conductive surface of the ITO conductive glass, and naturally airing at room temperature;
3) continuously dropwise adding 6 muL and 4 mg/mL zinc-doped cadmium sulfide aqueous solution on the surface of the modified electrode, and naturally airing;
4) continuously dropwise adding 6 mu L of gold nanoparticle aqueous solution on the surface of the modified electrode, and naturally airing at room temperature;
5) dropwise adding 4 muL of procalcitonin antibody of 10 mug/mL, washing with ultrapure water, and naturally airing at room temperature to a wet film state;
6) dropwise adding a bovine serum albumin solution with the mass fraction of 0.2% and prepared by using a PBS buffer solution with the pH of 7.0 to the surface of the modified electrode, washing the surface of the electrode by using ultrapure water, and airing in a refrigerator at 4 ℃;
7) dropwise adding 6 mu L of procalcitonin antigen, using 0.1 pg/mL-1000 ng/mL of PBS buffer solution for preparation, washing the surface of the electrode with ultrapure water, and naturally airing in a refrigerator at 4 ℃ to prepare the photoelectric chemical sensor for detecting the procalcitonin antigen.
EXAMPLE 3 preparation of photoelectrochemical sensor
(1) Preparation of bismuth molybdate nanosheet material
Dissolving 2.0g of pentahydrate bismuth nitrate and 0.6 g of sodium molybdate dihydrate in 150 mL of ultrapure water under stirring, stirring the solution at room temperature for 3 h, transferring the obtained light yellow solution after uniform stirring into a high-pressure reaction kettle, reacting at 200 ℃ for 24 h, naturally cooling to room temperature after the reaction is finished, washing the product with ultrapure water and absolute ethyl alcohol for 3 times respectively, and then vacuum-drying at 50 ℃ for 12 h to obtain the flaky bismuth molybdate nano material;
(2) preparation of zinc-doped cadmium sulfide nanoparticles
Proportionally dissolving zinc acetate dihydrate and cadmium acetate dihydrate in 50 mL of ultrapure water to ensure that the molar mass of zinc ions and cadmium ions is 0.01 mol in total, and uniformly stirring the solution at room temperature to obtain a solution A; dissolving 3.0g of sodium sulfide nonahydrate in 50 mL of ultrapure water, and uniformly stirring to obtain a solution B; mixing the solution A and the solution B, stirring at room temperature for 12 h, then pouring the mixed aqueous solution into a high-pressure reaction kettle, reacting at 200 ℃ for 18 h, naturally cooling to room temperature after the reaction is finished, washing the product for 3 times by using ultrapure water and absolute ethyl alcohol respectively, and then carrying out vacuum drying at 50 ℃ overnight to obtain the zinc-doped cadmium sulfide nano-particle material;
(3) preparation of gold nanoparticles
Boiling 50 mL of chloroauric acid aqueous solution with the mass fraction of 0.05% at 120 ℃ for 10 min, then adding 5.0 mL of trisodium citrate aqueous solution with the mass fraction of 3% into the boiled solution, continuously stirring the solution under the boiling condition, changing the color of the solution from slight blue to bright red, and continuously stirring to obtain purple solution, thus obtaining gold nanoparticles;
(4) preparation of photoelectrochemical sensor
1) Ultrasonically cleaning conductive glass by using a detergent, acetone, ethanol and ultrapure water in sequence, and drying by using nitrogen;
2) dripping 6 muL and 6 mg/mL aqueous solution of bismuth molybdate to the conductive surface of the ITO conductive glass, and naturally airing at room temperature;
3) continuously dropwise adding 6 muL and 6 mg/mL zinc-doped cadmium sulfide aqueous solution on the surface of the modified electrode, and naturally airing;
4) continuously dropwise adding 6 mu L of gold nanoparticle aqueous solution on the surface of the modified electrode, and naturally airing at room temperature;
5) dropwise adding 4 muL of procalcitonin antibody of 20 mug/mL, washing with ultrapure water, and naturally airing at room temperature to a wet film state;
6) dropwise adding a bovine serum albumin solution with the mass fraction of 0.3% and prepared by using a PBS buffer solution with the pH of 7.0 to the surface of the modified electrode, washing the surface of the electrode by using ultrapure water, and airing in a refrigerator at 4 ℃;
7) dropwise adding 6 mu L of procalcitonin antigen, using 0.1 pg/mL-1000 ng/mL of PBS buffer solution for preparation, washing the surface of the electrode with ultrapure water, and naturally airing in a refrigerator at 4 ℃ to prepare the photoelectric chemical sensor for detecting the procalcitonin antigen.
Example 4 detection of Procalcitonin
(1) An electrochemical workstation is used for testing by a three-electrode system, a saturated calomel electrode is used as a reference electrode, a platinum wire electrode is used as an auxiliary electrode, the prepared ITO modified sensor is used as a working electrode, and the test is carried out in 10 mL of PBS (phosphate buffer solution) containing 0.01 mol/L ascorbic acid and having the pH value of 5.0;
(2) detecting procalcitonin antigen by a time-current method, setting the voltage to be-0.1V, the running time to be 120s and the light source wavelength to be 400 nm;
(3) after the electrodes are placed, turning on the lamp every 20s for continuously irradiating for 20s, recording the photocurrent, and drawing a working curve;
(4) and replacing the procalcitonin antigen sample solution to be detected with the procalcitonin antigen standard solution for detection.
Example 5 detection of Procalcitonin
(1) Testing by using an electrochemical workstation in a three-electrode system, taking a saturated calomel electrode as a reference electrode, taking a platinum wire electrode as an auxiliary electrode, taking the prepared ITO modified sensor as a working electrode, and testing in 10 mL of PBS (phosphate buffer solution) containing 0.1 mol/L ascorbic acid and having the pH value of 7.0;
(2) detecting procalcitonin antigen by a time-current method, setting the voltage to be 0V, the running time to be 120s and the light source wavelength to be 430 nm;
(3) after the electrodes are placed, turning on the lamp every 20s for continuously irradiating for 20s, recording the photocurrent, and drawing a working curve;
(4) and replacing the procalcitonin antigen sample solution to be detected with the procalcitonin antigen standard solution for detection.
Example 6 detection of Procalcitonin
(1) Testing by using an electrochemical workstation in a three-electrode system, taking a saturated calomel electrode as a reference electrode, taking a platinum wire electrode as an auxiliary electrode, taking the prepared ITO modified sensor as a working electrode, and testing in 10 mL of PBS (phosphate buffer solution) containing 0.5 mol/L ascorbic acid and having the pH value of 8.0;
(2) detecting procalcitonin antigen by a time-current method, setting the voltage to be 0.1V, the running time to be 120s and the light source wavelength to be 450 nm;
(3) after the electrodes are placed, turning on the lamp every 20s for continuously irradiating for 20s, recording the photocurrent, and drawing a working curve;
(4) and replacing the procalcitonin antigen sample solution to be detected with the procalcitonin antigen standard solution for detection.
Claims (1)
1. A preparation method of a photoelectrochemical immunosensor based on bismuth molybdate/zinc-doped cadmium sulfide/gold is characterized by comprising the following steps:
(1) preparation of bismuth molybdate nanosheet material
Dissolving 1.0-2.0 g of pentahydrate bismuth nitrate and 0.4-0.6 g of sodium molybdate dihydrate into 100-150 mL of ultrapure water under stirring, stirring the solution at room temperature for 1-3 h, transferring the uniformly stirred pale yellow solution into a high-pressure reaction kettle, reacting at 160-200 ℃ for 20-24 h, naturally cooling to room temperature after the reaction is finished, washing the obtained product with ultrapure water and absolute ethyl alcohol for 3 times respectively, and then vacuum-drying at 30-50 ℃ for 8-12 h to prepare a bismuth molybdate nanosheet material;
(2) preparation of zinc-doped cadmium sulfide nanoparticles
Taking zinc acetate dihydrate and cadmium acetate dihydrate according to different proportions, dissolving the reagents into 20-50 mL of ultrapure water together to ensure that the molar mass of zinc ions and cadmium ions is 0.01 mol in total, and uniformly stirring the solution at room temperature to obtain a solution A; dissolving 1.0-3.0 g of sodium sulfide nonahydrate in 20-50 mL of ultrapure water, and uniformly stirring to obtain a solution B; mixing the solution A and the solution B, stirring for 6-12 h at room temperature, then pouring the mixed aqueous solution into a high-pressure reaction kettle, reacting for 14-18 h at 150-200 ℃, naturally cooling to room temperature after the reaction is finished, washing the obtained product for 3 times by using ultrapure water and absolute ethyl alcohol respectively, and then carrying out vacuum drying at 30-50 ℃ overnight to obtain a zinc-doped cadmium sulfide nano-particle material;
(3) preparation of gold nanoparticles
Boiling 30-50 mL of chloroauric acid aqueous solution with the mass fraction of 0.01-0.05% at 120 ℃ for 5-10 min, then adding 2.0-5.0 mL of trisodium citrate aqueous solution with the mass fraction of 1-3% into the boiled solution, continuously stirring the solution under the boiling condition, changing the color of the solution from slight blue to bright red, and continuously stirring to obtain a purple solution, thus preparing gold nanoparticles;
(4) preparation of photoelectrochemical immunosensor
1) Ultrasonically cleaning conductive glass by using a detergent, acetone, ethanol and ultrapure water in sequence, and drying by using nitrogen;
2) dripping 6 muL and 2-6 mg/mL bismuth molybdate water solution onto a conductive surface of the ITO conductive glass, and naturally airing at room temperature;
3) continuously dropwise adding zinc-doped cadmium sulfide aqueous solution of 6 muL and 2-6 mg/mL on the surface of the modified electrode, and naturally airing;
4) continuously dropwise adding 6 mu L of gold nanoparticle aqueous solution on the surface of the modified electrode, and naturally airing at room temperature;
5) dropwise adding 4 muL of procalcitonin antibody of 5-20 mug/mL, washing with ultrapure water, and naturally airing at room temperature to a wet film state;
6) dropwise adding a bovine serum albumin solution with the mass fraction of 0.1% -0.3% and prepared by using a PBS buffer solution with the pH of 7.0 to the surface of the modified electrode, washing the surface of the electrode by using ultrapure water, and airing in a refrigerator at 4 ℃;
7) dropwise adding 6 mu L of procalcitonin antigen, using 0.1 pg/mL-1000 ng/mL of PBS buffer solution for preparation, washing the surface of the electrode with ultrapure water, and naturally airing in a refrigerator at 4 ℃ to prepare the photoelectrochemical immunosensor for detecting the procalcitonin antigen.
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