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Alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate radical

A technology of alpha-fetoprotein and detection method, which is applied in the field of detection and analysis, can solve the problems of complex instruments, etc., and achieve the effects of less reagent consumption, simple and instant detection, and low cost

Active Publication Date: 2022-05-06
SHANDONG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] Therefore, the clinical molecular diagnosis of AFP is very important. The currently reported immunoassay methods for the determination of AFP include fluorescence immunoassay, electrochemical immunoassay, chemiluminescence immunoassay and radioimmunoassay, although these methods have little effect on the sensitive detection of biomarkers. made a great contribution, but complex instruments and skilled professional operators must be provided and acquired

Method used

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  • Alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate radical
  • Alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate radical
  • Alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate radical

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Embodiment 1

[0046] Determination of copper ion concentration in embodiment 1 copper alginate hydrogel

[0047] Sodium alginate was dissolved in ultrapure water to prepare a 0.2 wt% sodium alginate solution. Dissolve copper sulfate in ultrapure water to prepare gradient copper sulfate solutions with concentrations of 0.5mM, 1mM, 1.5mM, 2mM and 2.5mM, respectively. 100 μL of sodium alginate solution and 100 μL of gradient copper sulfate solution were mixed uniformly at room temperature to obtain copper alginate hydrogel. After 2 hours, 35 μL of copper alginate hydrogel was added dropwise to one end of the pH test strip. After 3 minutes, the copper alginate hydrogel crawled on the pH test strip, and the moving distance was measured. The results are as follows: figure 2 shown.

[0048] Depend on figure 2 It can be seen that when the concentration of copper ions is less than 1mM, the formed mixture has a smaller viscosity and a longer distance to crawl on the test strip. The crawl distan...

Embodiment 2

[0049] The determination of embodiment 2 sodium pyrophosphate concentration

[0050] Sodium pyrophosphate was dissolved in 10 mM HEPES buffer solution, adjusted to pH = 7.4, and prepared into gradient sodium pyrophosphate solutions with concentrations of 0.5 mM, 1 mM, 1.5 mM and 2 mM respectively. Add 200 μL of the gradient sodium pyrophosphate solution to 200 μL of the copper alginate hydrogel prepared in Example 1, and mix well at room temperature by vortexing to obtain the pyrophosphate-copper alginate hydrogel. After 2 hours, 35 μL of pyrophosphate-copper alginate hydrogel was added dropwise to one end of the pH test paper strip. After 3 minutes, the pyrophosphate-copper alginate hydrogel crawled on the pH test paper strip, and the moving time was measured. distance, the result is image 3 shown.

[0051] Due to the strong coordination between pyrophosphate and copper ions, it will rob copper ions in the hydrogel. As the concentration of pyrophosphate increases, the deg...

Embodiment 3

[0052] The detection of embodiment 3 alkaline phosphatase

[0053] A kind of alkaline phosphatase detection method based on copper ion response pyrophosphate, comprises steps as follows:

[0054] Dissolve alkaline phosphatase in 10mM HEPES buffer solution (pH=7.4) and prepare gradient alkaline with concentrations of 1mU / mL, 10mU / mL, 25mU / mL, 50mU / mL, 75mU / mL, 100mU / mL Phosphatase solution. Add 200 μL of gradient alkaline phosphatase solution to 200 μL of 1.5 mM pyrophosphate solution, and incubate at 37°C for 60 min. Each reaction mixture was added to 200 μL of the copper alginate hydrogel prepared in Example 1, and mixed well at room temperature by vortexing to obtain an alkaline phosphatase mixture. After 2 hours, 35 μL of the alkaline phosphatase mixture was added dropwise to one end of the pH test strip. After 3 minutes, the alkaline phosphatase mixture crawled on the pH test strip, and the moving distance was measured. The results were as follows: Figure 4 shown. The...

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Abstract

The invention relates to an alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate. Comprising the following steps: (1) preparing a copper alginate hydrogel and a sodium pyrophosphate solution; (2) preparing an alkaline phosphatase standard solution and an alpha fetoprotein standard solution with gradient concentrations; (3) drawing an alkaline phosphatase standard curve; (4) drawing an alpha fetoprotein standard curve; and (5) detecting the contents of alkaline phosphatase and alpha fetoprotein in the sample to be detected. The alkaline phosphatase or alpha fetoprotein detection method based on copper ion response pyrophosphate radical provided by the invention has the advantages of simple operation process, low cost, low reagent consumption and the like, the detection limit of alkaline phosphatase is 1.7 mU / mL, the quantitative detection limit of alpha fetoprotein is 0.8 ng / mL, the problems of complexity and high cost of the existing detection method are effectively solved, and the method is suitable for industrial production. And a development prospect is provided for clinical diagnosis and application.

Description

technical field [0001] The invention provides a method for detecting alkaline phosphatase or alpha-fetoprotein based on copper ion response to pyrophosphate, and belongs to the technical field of detection and analysis. Background technique [0002] Alkaline phosphatase (ALP) is one of the most important hydrolytic enzymes widely distributed in animals. It can catalyze the dephosphorylation of biomolecules and regulate cell cycle processes such as cell growth, apoptosis, and signal transduction. In clinical diagnosis, ALP as an important biomarker can indicate a variety of diseases, such as diabetes, bone disease, prostate cancer and abnormal liver function. In addition, ALP is widely used as a labeled enzyme in enzyme-linked immunosorbent assay (ELISA) due to its high catalytic activity, good stability, wide substrate specificity, and easy binding to antibodies. The highly specific antigen-antibody recognition and efficient biocatalytic properties of labeled enzymes make E...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/58C12Q1/42
CPCG01N33/57476G01N33/581C12Q1/42G01N2333/916
Inventor 于丽台文君宓静如吴达刘金鹏
Owner SHANDONG UNIV
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