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Measurement method of acid phosphatase activity

An acid phosphatase and assay method technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of poor anti-interference ability, cumbersome detection process, low sensitivity, etc., and achieves strong anti-interference ability and sensitivity. The effect of high and high sensitivity detection

Inactive Publication Date: 2016-05-04
NANJING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above-mentioned methods all have the disadvantages of low sensitivity, poor anti-interference ability, cumbersome detection process, etc.

Method used

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  • Measurement method of acid phosphatase activity
  • Measurement method of acid phosphatase activity
  • Measurement method of acid phosphatase activity

Examples

Experimental program
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Effect test

Embodiment 1

[0026] Example 1: Feasibility analysis of colorimetric analysis of acid phosphatase activity by the method of the present invention.

[0027] 20 μL acid phosphatase solution (100mU·mL -1 ) with 20 μL-ascorbic acid-2-phosphate (0.3M), 10 μL copper sulfate-Yulketonin disulfonate complex (copper sulfate: 12 mM, yloketone sodium disulfonate: 24 mM) and 550 μL glycine-HCl buffer (50mM, pH5.7) were evenly mixed, and the resulting mixture was incubated at 25°C for 15min, then the UV-Vis absorption spectrum of the solution in the range of 800-300nm was measured and the color change of the solution was recorded with a digital camera, as figure 2 shown. Among them, A means that all reagents are added, B means that acid phosphatase is not added, C means that L-ascorbic acid-2-phosphate is not added, and D means that copper sulfate-bathone sodium disulfonate complex is not added. The reagents were replaced with an equal amount of 50 mM glycine-hydrochloric acid buffer at pH 5.7. From ...

Embodiment 2

[0028] Example 2: The relationship between the ultraviolet-visible absorption spectrum and color change of the detection solution and the activity of acid phosphatase.

[0029] 20 μL of 0, 10, 20, 30, 40, 50, 60, 70, 80, 90 and 100 mU·mL -1 The acid phosphatase solution was mixed with 20 μL-ascorbic acid-2-phosphate (0.3M), 10 μL copper sulfate-thyroxine sodium disulfonate complex (copper sulfate: 12 mM, yloxone disulfonate sodium: 24 mM) and 550 μL of glycine-hydrochloric acid buffer solution (50 mM, pH5.7) was evenly mixed, and the resulting mixture was incubated at 25°C for 15 min, and then the UV-visible absorption spectrum of the solution in the range of 800-300 nm was measured and recorded with a digital camera The color changes, such as image 3 shown. From image 3 (A) It can be seen that with the increase of acid phosphatase activity in the sample, the absorption value of the detection solution at 484nm increases correspondingly. From image 3 (B) It can be seen ...

Embodiment 3

[0030] Example 3: Selective analysis of the colorimetric analysis of acid phosphatase activity by the method of the present invention.

[0031]20μL 1mg·mL -1 Acid Phosphatase (A), Bovine Serum Albumin (B), Hemoglobin (C), Horseradish Peroxidase (D), Streptavidin (E), Thrombin (F), Trypsin (G) Or 50mM pH5.7 glycine-hydrochloric acid buffer (H) was mixed with 20μL-ascorbic acid-2-phosphate (0.3M), 10μL copper sulfate-thyroxine sodium disulfonate complex (copper sulfate: 12mM, yloxone Sodium disulfonate: 24mM) and 550μL of glycine-hydrochloric acid buffer (50mM, pH5.7) were uniformly mixed, and the resulting mixture was incubated at 25°C for 15min, then the absorbance of the solution at 484nm was measured and recorded with a digital camera Record the color change of the solution as Figure 4 shown. It can be seen from the figure that except acid phosphatase, other interfering proteins will not cause obvious changes in the absorption value or color of the detection solution at ...

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Abstract

The invention discloses a measurement method of acid phosphatase activity. According to the method, L-ascorbic acid-2-phosphoric acid serves as a catalytic substrate of acid phosphatase, the substrate is hydrolyzed into L-ascorbic acid under catalysis of acid phosphatase, because bivalent copper can be reduced into monovalent copper by L-ascorbic acid so that a bivalent copper compound serving as a color developing probe can be converted into a monovalent copper compound in deep color, the higher the activity of acid phosphatase in samples is, the more obvious the color change is, and therefore semi-quantitative colorimetric analysis of the activity of acid phosphatase is achieved, or quantitative detection can be performed with an ultraviolet-visible spectrophotometer. By means of the method, acid phosphatase with the concentration of 0.48 mU.mL<-1> can be detected. The method is high in sensitivity, good in selectivity, high in anti-interference capacity, good in repeatability, easy to operate, short in detection time and low in cost, can be used for quantitatively measuring the activity of acid phosphatase in serum samples, and a fast, simple, convenient, low-cost, sensitive and stable-performance colorimetric analysis method is provided for detecting the activity of acid phosphatase in clinical samples.

Description

technical field [0001] The invention belongs to the technical field of biological analysis, in particular to a method for measuring acid phosphatase activity, in particular to a method for quickly measuring acid phosphatase activity based on colorimetric analysis. Background technique [0002] Acid phosphatase (acid phosphatase, EC3.1.3.2, ACP) mainly exists in macrophages, localized in lysosomes, and can catalyze the hydrolysis of phosphate monoesters to produce inorganic phosphoric acid and corresponding alcohols, phenols or sugars. The ACP in normal human serum mainly comes from tissues such as prostate, bone, liver, kidney, spleen, and pancreas, and its content is roughly the same regardless of men, women, or children. And patients with prostate disease and osteomyelitis deformans, osteogenesis imperfecta, rickets, osteosarcoma, multiple myeloma, primary bone tumors, hyperthyroidism, leukemia, breast cancer, myocardial infarction, hepatitis, liver cirrhosis, gallbladder ...

Claims

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Application Information

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IPC IPC(8): C12Q1/42
CPCC12Q1/42
Inventor 孔金明胡琼梅亚琦何玟辉张学记
Owner NANJING UNIV OF SCI & TECH
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