CN114674891B - Construction of sensor with hollow structure combined with electron consumption strategy - Google Patents

Abstract

The invention discloses a construction method of a sensor with a hollow structure combined with an electron consumption strategy. Firstly, a hollow structure ZnCdS/ZnIn ₂ S ₄ heterojunction is synthesized. The hierarchical hollow structure of the heterojunction can promote light collection through multiple light scattering/reflection, At the same time, the synergy between the hollow structure and the close contact heterojunction interface can promote visible light harvesting, accelerate charge transfer and inhibit the recombination of photogenerated carriers; in addition, AuNPs‑AAO is modified on the secondary antibody as a signal amplification carrier, AAO Under this effect, AAO catalyzes the oxidation of AA, leading to a reduction in the amount of AA, thereby greatly reducing the photocurrent response.

Description

Construction of sensor with hollow structure combined with electron consumption strategy

technical field

The invention relates to the field of quantitative detection of prostate specific antigen, more specifically to the construction of a sensor with a hollow structure combined with an electron consumption strategy.

Background technique

Prostate cancer is a malignant epithelial tumor of the prostate gland that is common in men worldwide. Prostate lesions are asymptomatic in the early stage and can be fatal in severe cases. Therefore, the realization of early detection and treatment of prostate cancer plays a very important role in the diagnosis of malignant tumors. The establishment of a simple, rapid, sensitive and selective method for the detection of malignant tumor biomarkers has a great impact on the early detection and treatment of malignant tumors. Evaluation is of great value.

Photoelectrochemical (PEC) immunosensors have attracted much attention due to their advantages such as low background signal, high sensitivity, simple operation, and inexpensive instruments. This is a sensitive detection technology combining photoelectrochemistry and immunology, which has both the strong specificity of immunology and the high sensitivity of photoelectrochemistry. Simply put, the detection mechanism of the PEC immunosensor is to convert the content of the detected substance into a photoelectric signal through signal amplification under light conditions.

Contents of the invention

The purpose of the present invention is to construct a photoelectrochemical sensor for prostate-specific antigen detection by using a hollow structure combined with an electron consumption strategy sensor.

In order to solve the above-mentioned technical problems, the present invention is achieved through the following measures: the construction of a sensor with a hollow structure combined with an electronic consumption strategy, which is characterized by the following steps:

(1) Synthesis of ZIF-8: 2.36 g of Zn(NO ₃ ) ₂ ·6H ₂ O was dissolved in 200 mL of methanol, and 1.32 g of 2-methylimidazole was quickly added to the above solution under stirring, followed by Aged at room temperature for 11 h; finally, washed with ethanol three times and dried in vacuum at 70 °C for 12 h.

(2) Synthesis of ZIF-8-derived hollow ZnCdS: Mix 50 mL of ethanol solution with 20 mg of ZIF-8 synthesized in step (1) and 400 mg of thioacetamide, transfer to a 100 mL autoclave, and keep at 150 °C After 1 h, the product was collected by centrifugation, and washed with ethanol to obtain a purple product; thereafter, 20 mg of the purple product and 78 _mg of CdCl were dissolved in 25 mL of ethanol, and after magnetic stirring, the mixture was transferred to a 50 mL autoclave and heated at 160 After heating at ℃ for 4 h, the obtained yellow precipitate was collected by centrifugation, washed with ethanol, and then dried in vacuum at 70 °C for 12 h.

(3) Synthesis of hollow ZnCdS/ZnIn ₂ S ₄ heterojunction: Weigh 0.2 g of the powder obtained in step (2) and disperse it in 20 mL of ethanol, sonicate for 30 min; then, mix 1 mmol ZnCl ₂ , 2 mmol InCl ₃ · 4H ₂ O and 8 mmol thioacetamide were added to the above suspension and stirred vigorously for 30 min; the resulting mixture was sealed in a 50 mL polytetrafluoroethylene-lined autoclave and kept at 120 °C for 2 h; The product was extracted by centrifugation, washed with deionized water and ethanol, and dried at 60 °C.

(4) Synthesis of ITO/ZnCdS/ZnIn ₂ S ₄ electrodes: The conductive glass is indium tin oxide glass (ITO). Cut the conductive glass into 4.0×0.5 cm strips, and use acetone solution, double distilled water and absolute ethanol in sequence Ultrasonic cleaning for 5 min, and then drying under nitrogen for use; the concentration of 2.0 mg/mL ZnCdS/ZnIn ₂ S ₄ synthesized in step (3) was added dropwise on the ITO glass, and dried at 60°C to obtain ITO/ZnCdS/ZnIn ₂ S ₄ electrodes.

(5) Synthesis of AuNPs: under continuous stirring, 1.5 mL of 25 mmol/L HAuCl ₄ and 5 mL of 10 mmol/L sodium citrate were mixed in a round bottom flask; after that, 1.5 mL of 0.1 mol/L NaBH ₄ was quickly injected into the above solution, and then it turned orange-red, indicating the formation of particles; thereafter, the mixed solution was stirred at room temperature for another 6 h, and the color changed from orange-red to wine red; finally, the resulting AuNPs solution was heated at 4 ° Store under C.

(6) Synthesis of AuNPs-AAO: First, add 0.2 mol K ₂ CO ₃ dropwise, and stir to adjust the pH of the Au NPs solution synthesized in step (5) to 8.2; then, add 40 μL of 0.8 mg/mL ascorbate oxidase (AAO) They were added to 2.0 mL alkaline AuNPs solution, incubated for 2 h, the solution mixture was centrifuged at 10000 rpm for 20 min, and washed several times to remove residues.

(7) Synthesis of Ab2-AuNPs-AAO: Take 1 mL of the secondary antibody with a concentration of 10 μg/mL, that is, Ab2, and add it to the product synthesized in step (6), incubate at 4 °C for 2 h, and use pH 7.4 phosphate Ab2 without complexation was removed by buffer washing three times, and Ab2-AuNPs-AAO was obtained.

(8) Construction of photoelectric sensor (PEC): Rinse the ITO/ZnCdS/ZnIn ₂ S ₄ electrode with ultrapure water, then incubate 6 μL of the primary antibody (Ab1) with a concentration of 10 μg/mL at 4 ºC for 16 h, and use Wash thoroughly with pH 7.4 phosphate buffer 3 times; continue to drop-coat 20 µL of 3% bovine serum albumin to block non-specific binding sites, wash thoroughly with pH 7.4 phosphate buffer 3 times, add 20 µL of different concentrations Prostate antigen was added dropwise to the surface of the electrode, incubated at room temperature for 30 min, and washed three times with pH 7.4 phosphate buffer; continued to drop 20 μL of Ab2-AuNPs-AAO synthesized in step (7), and incubated at room temperature for 4 h.

(9) Electrochemical detection of photoelectric sensors: the modified electrode processed in step (7) is used as the working electrode, the counter electrode is a platinum wire electrode, the reference electrode is an Ag/AgCl electrode, the bias value is 0 V, and the xenon lamp is used as a light source Stimulation, the electrolytic cell is a phosphate buffer system with pH 7.4 (containing 1 mol/L ascorbic acid), and the current I-T curve is measured to detect the photoelectric performance.

Beneficial effects of the present invention:

(1) The present invention has low cost, simple experimental operation, and easy control of reaction conditions.

(2) ZnCdS/ZnIn ₂ S ₄ heterojunction hierarchical hollow structure can facilitate light harvesting through multiple light scattering/reflection.

(3) The synergy between the ZnCdS/ZnIn ₂ S ₄ heterojunction hollow structure and the close-contact heterojunction interface, which can promote visible light harvesting, accelerate charge transfer, and suppress the recombination of photogenerated carriers.

(4) AAO-induced quenching effect to suppress the PEC signal, under this effect, AAO catalyzes the oxidation of AA, leading to a decrease in the amount of AA, thereby greatly reducing the photocurrent response.

Detailed ways

In order to further understand the present invention, carry out according to the technical scheme of the present invention in conjunction with embodiment, provide specific implementation mode:

(1) Synthesis of ZIF-8: 2.36 g of Zn(NO ₃ ) ₂ ·6H ₂ O was dissolved in 200 mL of methanol, and 1.32 g of 2-methylimidazole was quickly added to the above solution under stirring, followed by Aged at room temperature for 11 h; finally, washed with ethanol three times and dried in vacuum at 70 °C for 12 h.

(2) Synthesis of ZIF-8-derived hollow ZnCdS: Mix 50 mL of ethanol solution with 20 mg of ZIF-8 synthesized in step (1) and 400 mg of thioacetamide, transfer to a 100 mL autoclave, and keep at 150 °C After 1 h, the product was collected by centrifugation, and washed with ethanol to obtain a purple product; thereafter, 20 mg of the purple product and 78 _mg of CdCl were dissolved in 25 mL of ethanol, and after magnetic stirring, the mixture was transferred to a 50 mL autoclave and heated at 160 After heating at ℃ for 4 h, the obtained yellow precipitate was collected by centrifugation, washed with ethanol, and then dried in vacuum at 70 °C for 12 h.

(3) Synthesis of hollow ZnCdS/ZnIn ₂ S ₄ heterojunction: Weigh 0.2 g of the powder obtained in step (2) and disperse it in 20 mL of ethanol, sonicate for 30 min; then, mix 1 mmol ZnCl ₂ , 2 mmol InCl ₃ · 4H ₂ O and 8 mmol thioacetamide were added to the above suspension and stirred vigorously for 30 min; the resulting mixture was sealed in a 50 mL polytetrafluoroethylene-lined autoclave and kept at 120 °C for 2 h; The product was extracted by centrifugation, washed with deionized water and ethanol, and dried at 60 °C.

(4) Synthesis of ITO/ZnCdS/ZnIn ₂ S ₄ electrodes: The conductive glass is indium tin oxide glass (ITO). Cut the conductive glass into 4.0×0.5 cm strips, and use acetone solution, double distilled water and absolute ethanol in sequence Ultrasonic cleaning for 5 min, and then drying under nitrogen for use; the concentration of 2.0 mg/mL ZnCdS/ZnIn ₂ S ₄ synthesized in step (3) was added dropwise on the ITO glass, and dried at 60°C to obtain ITO/ZnCdS/ZnIn ₂ S ₄ electrodes.

(5) Synthesis of AuNPs: 1.5 mL of 25 mmol/L HAuCl ₄ and 5 mL of 10 mmol/L sodium citrate were mixed in a round bottom flask under continuous stirring; after that, 1.5 mL of 0.1 mol /L of NaBH ₄ was quickly injected into the above solution, and then it turned orange-red, indicating the formation of particles; thereafter, the mixed solution was stirred at room temperature for another 6 h, and the color changed from orange-red to wine red; finally, the generated AuNPs solution was Store at 4°C.

(6) Synthesis of AuNPs-AAO: First, add 0.2 mol K ₂ CO ₃ dropwise, and stir to adjust the pH of the Au NPs solution synthesized in step (5) to 8.2; then, add 40 μL of 0.8 mg/mL ascorbate oxidase (AAO) They were added to 2.0 mL alkaline AuNPs solution, incubated for 2 h, the solution mixture was centrifuged at 10000 rpm for 20 min, and washed several times to remove residues.

(7) Synthesis of Ab2-AuNPs-AAO: Take 1 mL of the secondary antibody with a concentration of 10 μg/mL, that is, Ab2, and add it to the product synthesized in step (6), incubate at 4 °C for 2 h, and use pH 7.4 phosphate Ab2 without complexation was removed by buffer washing three times, and Ab2-AuNPs-AAO was obtained.

(8) Construction of photoelectric sensor (PEC): Rinse the ITO/ZnCdS/ZnIn ₂ S ₄ electrode with ultrapure water, then incubate 6 μL of the primary antibody (Ab1) with a concentration of 10 μg/mL at 4 ºC for 16 h, and use Wash thoroughly with pH 7.4 phosphate buffer 3 times; continue to drop-coat 20 µL of 3% bovine serum albumin to block non-specific binding sites, wash thoroughly with pH 7.4 phosphate buffer 3 times, add 20 µL of different concentrations Prostate antigen was added dropwise to the electrode surface, incubated at room temperature for 30 min, and washed three times with pH 7.4 phosphate buffer; continued to drop 20 μL of Ab2-AuNPs-AAO synthesized in step (7), and incubated at room temperature for 4 h.

(9) Electrochemical detection of photoelectric sensors: the modified electrode processed in step (7) is used as the working electrode, the counter electrode is a platinum wire electrode, the reference electrode is an Ag/AgCl electrode, the bias value is 0 V, and the xenon lamp is used as a light source Stimulation, the electrolytic cell is a phosphate buffer system of pH 7.4 (containing 1 mol/L ascorbic acid), the current I-T curve is measured to detect the photoelectric performance, and the linear equation is I=-1.04log(c)-10.44, The correlation coefficient is 0.992, and the detection limit is 0.04 pg/mL, realizing the detection of prostate specific antigen with high sensitivity.

Claims (1)

1. The construction method of the hollow structure combined with the electronic consumption strategy sensor is characterized by comprising the following steps:

(1) Synthesis of ZIF-8: zn (NO) of 2.36 g ₃ ) ₂ ·6H ₂ O was dissolved in 200 mL methanol and 1.32 g of 2-methylimidazole was added rapidly to the solution with stirring followed by aging at room temperature of 11 h; finally, washing 3 times with ethanol and vacuum drying 12 h at 70 ℃;

(2) Synthesis of ZIF-8 derived hollow ZnCdS: combining 50 mL ethanol solution with 20 mg ZIF-8 synthesized in step (1) and400 mixing mg of thioacetamide, transferring into a 100 mL autoclave, maintaining at 150 ℃ for 1 h, centrifuging and collecting a product, and washing with ethanol to obtain a purple product; thereafter 20 mg of the purple product and 78 mg of CdCl ₂ Dissolving in 25 mL ethanol, magnetically stirring, transferring the mixture into 50 mL autoclave, heating at 160deg.C for 4 h, centrifuging to collect yellow precipitate, washing with ethanol, and vacuum drying at 70deg.C for 12 h;

(3) Synthesis of hollow ZnCdS/ZnIn ₂ S ₄ Heterojunction: weighing 0.2 and g, dispersing the powder obtained in the step (2) in 20 mL ethanol, and carrying out ultrasonic treatment for 30 min; then, 1 mmol ZnCl ₂ 、2 mmol InCl ₃ ·4H ₂ O and 8 mmol thioacetamide were added to the above suspension and vigorously stirred for 30 min; the resulting mixture was sealed in a 50 mL teflon lined autoclave, held at 120 ℃ for 2 h; the product is centrifugally separated and extracted, washed by deionized water and ethanol and dried at 60 ℃;

(4) Construction of ITO/ZnCdS/ZnIn ₂ S ₄ An electrode: the conductive glass is indium tin oxide glass (ITO), the conductive glass is cut into 4.0x0.5 cm strips, sequentially washed by acetone solution, secondary distilled water and absolute ethyl alcohol for 5 min in an ultrasonic manner, and then dried under nitrogen for standby; znCdS/ZnIn synthesized in the step (3) with the concentration of 2.0 mg/mL ₂ S ₄ Dripping on ITO glass, drying at 60deg.C to obtain ITO/ZnCdS/ZnIn ₂ S ₄ An electrode;

(5) Synthesis of AuNPs: under continuous stirring, 1.5. 1.5 mL of HAuCl at a concentration of 25 mmol/L ₄ And 5 mL sodium citrate at a concentration of 10 mmol/L in a round bottom flask; thereafter, 0.1 mol/L NaBH of 1.5. 1.5 mL was added ₄ Rapidly infuse the solution before it turns orange-red indicating particle formation; thereafter, the mixed solution was stirred at room temperature for 6 h again, and the color was changed from orange red to reddish wine; finally, the AuNPs solution produced is stored at 4 ℃;

(6) Synthesis of AuNPs-AAO: first, 0.2 mol of K is added dropwise ₂ CO ₃ Stirring and adjusting the pH of the Au NPs solution synthesized in the step (5) to 8.2; subsequently, 40. Mu.L of 0.8. 0.8 mg/mL ascorbate oxidase (AAO)) Respectively adding into 2.0 mL alkaline Au NPs solution, culturing 2 h, centrifuging the solution mixture at 10000 rpm for 20 min, and cleaning for several times to remove residues;

(7) Synthesis of Ab2-AuNPs-AAO: adding the secondary antibody, namely Ab2, with the concentration of 1 mL of 10 mug/mL into the synthesized product of the step (6), incubating for 2 h at the temperature of 4 ℃, and washing for 3 times by using phosphate buffer solution with the pH of 7.4 to remove the Ab2 without complexing, thus obtaining Ab2-AuNPs-AAO;

(8) Construction of a Photosensor (PEC): rinsing ITO/ZnCdS/ZnIn with ultrapure water ₂ S ₄ The electrode was then incubated 16 h with 6 μl of primary antibody, ab1, at a concentration of 10 μg/mL at 4 ℃ and thoroughly rinsed 3 times with phosphate buffer pH 7.4; continuously dripping 20 mu L of 3% bovine serum albumin to block the non-specific binding site, thoroughly flushing 3 times by using phosphate buffer solution with the pH of 7.4, dripping 20 mu L of prostate antigens with different concentrations onto the surface of an electrode, incubating for 30 min at room temperature, and washing 3 times by using the phosphate buffer solution with the pH of 7.4; the 20. Mu.L of Ab2-AuNPs-AAO synthesized in step (7) was further added dropwise, and incubated at room temperature for 4 h.