A P-Ce based 0.3 Co 1.7 Preparation method of microcystin sensor based on alloy nanoflowers

Abstract

The present invention belongs to the technical field of preparation of sensing interfaces for detecting environmental pollutants such as microcystins, and specifically relates to a method for preparing a sensing interface based on phosphorus-doped cerium-cobalt (P‑Ce). 0.3 Co 1.7 Preparation method of electrode material of alloy nanoflower and application of electrode material. The preparation method of electrode material of microcystin based on signal amplification strategy of the present invention is to prepare P-Ce 0.3 Co 1.7 Alloy nanoflowers were used as the substrate material of the sensor to further prepare defect-rich ferrous sulfide coated bismuth oxide FeS 1‑x @Bi2O3 hybrid is used as a marker, and the excellent electrocatalytic activity of the base material and the marker material is utilized to apply the electrode material to ultrasensitive detection of microcystin. 0.3 Co 1.7 Preparation method of alloy nanoflower microcystin sensing platform, improving the detection signal strength and detection sensitivity of the sensor, and the detection range of microcystin is 0.01fg·mL ‑1 ~10ng·mL ‑1 , the detection limit was 4.2 fg·mL ‑1 The patent of this invention solves the problem that the traditional microcystin detection method cannot accurately detect low concentrations. This technology will provide a strong basis for future environmental analytical chemistry.

Description

Technical Field

[0001] The present invention belongs to the technical field of preparation of electrode materials for microcystin detection, and specifically relates to a method based on P-Ce 0.3 Co 1.7 Preparation method and application of electrode material of microcystin sensor based on alloy nanoflowers. Background Art

[0002] Microcystin is a cyclic heptapeptide compound produced during the outbreak of cyanobacteria blooms. It has strong hepatotoxicity. It can strongly inhibit the activity of cellular protein phosphatases 1 and 2A, impair their regulatory functions in the cell, and thus trigger primary liver cancer. In addition, microcystin can enter the food chain through drinking water or accumulation in aquatic products, and enter the human body directly or indirectly, posing a great threat to human health. In natural waters, microcystin is widely distributed and outbreaks are seasonal, so continuous monitoring and early warning are essential.

[0003] Electrochemical immunosensors are self-contained integrated devices based on antigen-antibody reactions that can perform specific quantitative or semi-quantitative analysis. In the detection of tumor markers, traditional technologies such as radioimmunoassay, enzyme immunoassay, chemiluminescence immunoassay and time-resolved fluorescence immunoassay have problems such as complex instruments and equipment, radioactive contamination, time-consuming analysis and expensive detection, which limit their clinical development. With the continuous development of cancer analysis technology, electrochemical immunosensors based on nanotechnology have attracted close attention from biological researchers due to their unique advantages. Electrochemical immunosensors use working electrodes as matrix platforms, and use biological molecules such as proteins, nucleic acids or antibodies as recognition elements to selectively react with target analytes, generating electrical response signals related to the concentration of target analytes, thereby achieving the purpose of quantitative detection of target analytes. Specifically, when antigens bind to antibodies, they cause changes in the charge distribution on the electrode surface, thereby generating electrical signals. This electrical signal can be measured and analyzed by equipment such as electrochemical workstations to obtain concentration information of the target analyte. Summary of the invention

[0004] Compared with the prior art, the present invention has the following beneficial effects: (1) This invention reports the synthesis of P-Ce for the first time 0.3 Co 1.7 Alloy nanoflowers have better conductivity and biocompatibility.

[0005] (2) The present invention prepares defect-rich ferrous sulfide coated bismuth oxide FeS 1-x @Bi2O3 hybrids are used as markers, and the defects produced by high-temperature calcination can accelerate the electrode interface reaction rate.

[0006] (3) The present invention constructs a P-Ce 0.3 Co 1.7 The microcystin sensor based on alloy nanoflowers has the advantages of low cost, high sensitivity, high selectivity, rapid detection, and easy preparation. It achieves the purpose of rapid and ultrasensitive detection of microcystin, and its detection linear range is 0.01 fg·mL -1 ~10 ng·mL -1 , the detection limit was 4.2 fg·mL -1 . BRIEF DESCRIPTION OF THE DRAWINGS

[0007] Figure 1 P-Ce prepared in step (1) of Example 1 0.3 Co 1.7 SEM image of alloy nanoflowers; Figure 2 P-Ce prepared in step (1) of Example 1 0.3 Co 1.7 Transmission electron microscopy image of alloy nanoflowers; Figure 3 FeS prepared in step (2) of Example 1 1-x @SEM image of Bi2O3 hybrid; Figure 4 This is a stability diagram of the microcystin sensor prepared in Example 2; Figure 5 This is a reproducibility graph of the microcystin sensor prepared in Example 2; Figure 6 This is a graph showing the linear range of detection of microcystin using the sensor prepared in Example 3. DETAILED DESCRIPTION

[0008] The present invention will be further described below in conjunction with specific embodiments. Example

[0009] The P-Ce 0.3 Co 1.7 The method for preparing the electrode material of the microcystin sensor of the alloy nanoflower comprises the following steps: (1) Preparation of P-Ce by electrodeposition 0.3 Co 1.7 Alloy Nanoflower Weigh Ce2(SO4)3 and CoSO4 in a ratio of (1:3) to (3:1) on an electronic balance, and prepare a solution with a total concentration of 0.05 mol / L with secondary water; weigh 0.01 mg of sodium phosphite in the above solution, sonicate the solution for 30 min at room temperature, and set aside; The volume of the electroplating solution was kept constant at 50 mL, and the pH of the solution was adjusted to 2 with a small amount of 1 mol / L sulfuric acid solution. The conventional three-electrode system was used to electro-deposit alloy nanoflowers, that is, Co sheet was used as the working electrode, Pt electrode was used as the counter electrode, and saturated Ag / AgCl electrode was used as the reference electrode. The constant potential polarization method was used for the experiment, and the potential was set to -0.8 V. The electro-deposition time was set to 100 s. After the electro-deposition was completed, the prepared electrode was thoroughly washed in secondary water to remove the residual plating solution components on the electrode surface, and then dried at room temperature to obtain phosphorus-doped cerium cobalt P-Ce grown on the Co sheet. 0.3 Co 1.7 / Co nanoflowers; (2) Defect-rich ferrous sulfide coated with bismuth oxide FeS 1-x Preparation of @Bi2O3 hybrids Weigh 20 mg of bismuth nitrate and dissolve it in 200 mL of secondary water. Heat the solution to 60°C and continuously add 5 mL of 1 mol / L sodium hydroxide aqueous solution to mix it. Heat the solution to 100°C and stir to dehydrate it into yellow bismuth trioxide Bi2O3. The obtained Bi2O3 was dispersed in 200 mL of secondary water, 10 mg of ferrous chloride and 10 mg of sodium sulfide were added, stirred rapidly, and the resulting precipitate was calcined at 300 °C for 3 h in a nitrogen atmosphere to obtain FeS 1-x @Bi2O3 hybrid; (3) Secondary antibody incubation Ab2-FeS 1-x Preparation of @Bi2O3 FeS 1-x @Bi2O3 and 3-aminopropyltriethoxysilane were dispersed in an ethanol solution, ultrasonically treated for 20 min, and then kept at 60-120 °C for 2 h. Amino-functionalized FeS was obtained by centrifugation, washing, and drying. 1-x @Bi2O3 nanospheres; Functionalize 10 mg of FeS with amino groups 1-x The Bi2O3 nanospheres were dispersed in 10 mL of ethanol-water solution, and 10 mL of phosphate buffer was added. The mixture was ultrasonicated for 20 min, and then 0.1 mL of microcystin antibody and 50 µL of glutaraldehyde were added. The mixture was stirred for 10 h, centrifuged and washed, and the secondary antibody Ab-FeS 1-x @Bi2O3. Example

[0010] The P-Ce 0.3 Co 1.7 The preparation method of the microcystin sensor of alloy nanoflowers comprises the following steps: (1) In P-Ce0.3 Co 1.7 10 μL of polydiallyldimethylammonium chloride was modified on the / Co electrode and dried at room temperature to form a film; (2) Add 6 µL and 1 µg / mL microcystin antibody solution Ab1 and 3 µL 0.5% BSA solution, wash with ultrapure water, and dry in a 4°C refrigerator; (3) Add 6 µL of a series of different concentrations of microcystin antigen solution Ag1, ranging from 0.01 to 5 ng / mL, rinse with ultrapure water, and dry in a refrigerator at 4°C; (4) Add 6 µL of secondary antibody Ab2-FeS 1-x @Bi2O3, rinse with ultrapure water, and dry in a 4°C refrigerator to obtain a sensor for microcystin. Example

[0011] The P-Ce 0.3 Co 1.7 The preparation method of the microcystin sensor of alloy nanoflowers, the detection of microcystin comprises the following steps: (1) The test was carried out using an electrochemical workstation with a three-electrode system, with the Ag / AgCl electrode as the reference electrode, the Pt electrode as the auxiliary electrode, and the prepared microcystin sensor as the working electrode. The test was carried out in 10 mL of a solution containing 4 mmol / L potassium ferrocyanide and 0.1 mmol / L potassium nitrate. (2) The tumor marker antigen standard solution was tested using square wave voltammetry, with the voltage test range being -0.2V to 0.6V; (3) When the background current tends to be stable, measure the peak current value of the sensor before and after the addition of the antigen, then record the current change and draw a working curve.

Claims

1. A P-Ce based 0.3 Co 1.7 The preparation method of the microcystin sensor of alloy nanoflowers and the preparation of the materials involved are characterized by: The following steps are involved: (1) Preparation of P-Ce by electrodeposition 0.3 Co 1.7 Alloy Nanoflower Weigh Ce2(SO4)3 and CoSO4 in a ratio of (1:3) to (3:1) on an electronic balance, and prepare a solution with a total concentration of 0.05 mol / L with double distilled water; weigh 0.01-0.5 mg of sodium phosphite in the above solution, sonicate the solution for 30 minutes at room temperature, and set aside; The volume of the electroplating solution was kept constant at 50 mL, and the pH of the solution was adjusted to 2-4 with a small amount of 1 mol / L sulfuric acid solution. The conventional three-electrode system was used to electro-deposit alloy nanoflowers, that is, Co sheet was used as the working electrode, Pt electrode was used as the counter electrode, and saturated Ag / AgCl electrode was used as the reference electrode. The constant potential polarization method was used for the experiment, and the potential was set to -0.8~-1.5V, and the electro-deposition time was set to 100~500 s. After the electro-deposition was completed, the prepared electrode was thoroughly washed in secondary water to remove the residual plating solution components on the electrode surface, and then dried at room temperature to obtain phosphorus-doped cerium cobalt P-Ce grown on the Co sheet. 0.3 Co 1.7 / Co nanoflowers; (2) Defect-rich ferrous sulfide coated with bismuth oxide FeS 1-x Preparation of @Bi2O3 hybrids Weigh 20-200 mg of bismuth nitrate and dissolve it in 200 mL of double distilled water. Heat the solution to 60-90 °C and continuously add 5-30 mL of 1 mol / L sodium hydroxide aqueous solution to mix it. Heat the solution to 100 °C and stir to dehydrate it into yellow bismuth trioxide Bi2O3. The obtained Bi2O3 was dispersed in 200 mL of secondary water, 10-200 mg of ferrous chloride and 10-100 mg of sodium sulfide were added, stirred rapidly, and the resulting precipitate was calcined at 300-550 °C for 3-10 h in a nitrogen atmosphere to obtain FeS 1-x @Bi2O3 hybrid; (3) Secondary antibody incubation Ab2-FeS 1-x Preparation of @Bi2O3 FeS 1-x @Bi2O3 and 3-aminopropyltriethoxysilane were dispersed in an ethanol solution, ultrasonically treated for 20-60 min, and then kept at 60-120 °C for 2-4 h. Amino-functionalized FeS was obtained by centrifugation, washing, and drying. 1-x @Bi2O3 nanospheres; Functionalize 10 mg of FeS with amino groups 1-x The Bi2O3 nanospheres were dispersed in 10-50 mL of ethanol-water solution, and 10-50 mL of phosphate buffer was added. The mixture was ultrasonicated for 20-30 min, and then 0.1 mL of microcystin antibody and 50 µL of glutaraldehyde were added. The mixture was stirred for 10-12 h, centrifuged and washed, and the secondary antibody Ab-FeS 1-x @Bi2O3.

2. A P-Ce based 0.3 Co 1.7 A method for preparing a microcystin sensor of alloy nanoflowers, wherein the sensor preparation method is characterized in that: The following steps are involved: (1) In P-Ce 0.3 Co 1.7 10~20 μL of polydiallyldimethylammonium chloride was modified on the / Co electrode and dried at room temperature to form a film; (2) Add 6~20 µL, 1~5 µg / mL microcystin antibody solution Ab1, and 3 µL of 0.5~2% BSA solution in sequence, wash with ultrapure water, and dry in a refrigerator at 4℃; (3) Add 6~20 μL of a series of different concentrations of microcystin antigen solution Ag1, 0.01~5 ng / mL, rinse with ultrapure water, and dry in a refrigerator at 4℃; (4) Add 6~20 µL of secondary antibody Ab2-FeS 1-x @Bi2O3, rinse with ultrapure water, and dry in a 4°C refrigerator to obtain a sensor for microcystin.

3. A P-Ce-based 0.3 Co 1.7 The preparation method of the microcystin sensor of alloy nanoflowers, the detection of microcystin, is characterized in that: The following steps are involved: (1) The test was carried out using an electrochemical workstation with a three-electrode system, with the Ag / AgCl electrode as the reference electrode, the Pt electrode as the auxiliary electrode, and the prepared microcystin sensor as the working electrode. The test was carried out in 10-20 mL of a solution containing 4-20 mmol / L potassium ferrocyanide and 0.1-10 mmol / L potassium nitrate. (2) The tumor marker antigen standard solution was tested using square wave voltammetry, with a voltage test range of -0.2V to 0.6V; (3) When the background current becomes stable, measure the peak current value of the sensor before and after the addition of the antigen, then record the current change and draw a working curve.

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