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Enzyme linked immunity diagnose reagent kit for HB core antigen detecting in two sandwich method and application thereof

A hepatitis B core antibody and diagnostic kit technology, which is applied in measurement devices, instruments, scientific instruments, etc., can solve the problem of difficulty in distinguishing weak positive samples from negative samples, and achieve high sensitivity, high specificity, and easy visual judgment. Effect

Inactive Publication Date: 2007-02-07
SHENYANG HUIMIN BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, traditional competitive inhibition assays are not easy to visually distinguish weakly positive samples from negative samples

Method used

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  • Enzyme linked immunity diagnose reagent kit for HB core antigen detecting in two sandwich method and application thereof
  • Enzyme linked immunity diagnose reagent kit for HB core antigen detecting in two sandwich method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Composition and Preparation of Double Sandwich ELISA Kit for Detecting Hepatitis B Core Antibody

[0024] Horseradish peroxidase: purchased from Faizyme;

[0025]Recombinant genetically engineered bacteria: E.coli JM105(pKK 223-3) ]; E.coli JM105(pFS 14NSD)[Florian Schodel et al.Structure of hepatitis B Virus Core ande-Antigen: a single precore amino acid prevents nucleocapsid assembly.J Biol Chem, 1993, 268(2): 1332~1337] ; E.coli M15 (pQE30) [Zhou Fuyuan et al., expressed and purified hepatitis B virus core antigen using pQE30 system. Chin J Biologicals, 2000, 13(1): 13-15];

[0026] Hepatitis B core antigen recombinant protein: Centrifuge the recombinant genetically engineered bacteria expressing hepatitis B core antigen at 8000rpm for 10 minutes to obtain bacterial pellet; resuspend the collected bacterial cells with 0.01mol / L pH7.4 phosphate buffer, ice bath, Bacteria for about 3 minutes until the suspension is clear; then centrifuge at 10,000 rpm for ...

Embodiment 2

[0038] Example 2 Detecting Hepatitis B Core Antibody Using Double Sandwich Method ELISA Kit

[0039] 1. Add 100 μl / well of the sample to be tested in the microwell reaction plate (the sample to be tested is diluted 1:30 with the sample diluent), set up two holes each for the negative control and positive control, and set up one hole for the blank control; seal the plate and place Incubate at 37°C for 1 hour;

[0040] 2. Wash the plate 3 times, 5 minutes each time

[0041] 3. Add 100 μl / well of enzyme-labeled antigen diluted 1:1200, and only add enzyme diluent to blank control wells; seal the plate and incubate at 37°C for 30 minutes;

[0042] 4. Wash the plate 3 times, 5 minutes each time

[0043] 5. Add substrate solution, seal the plate, and incubate at 37°C for 15 minutes;

[0044] 6. Add 50 μl / well stop solution to stop the reaction;

[0045] 7. Detection: Use an enzyme-labeled photometer to detect the light absorption value of each well at 450nm and 630nm; the detecti...

Embodiment 3

[0047] Example 3 Comparison of Double Sandwich ELISA Kit and Traditional Competitive Inhibition ELISA Kit for Detection of Hepatitis B Core Antibody

[0048] 1. We collected 943 cases of serum with different HBV status (Table 1). Each serum sample was detected simultaneously by double-antigen sandwich ELISA method and competitive inhibition ELISA method diagnostic kit (purchased from Shanghai Kehua Bioengineering Co., Ltd.). If the test result of the sample is marginal and there is a discrepancy between the two test results, use the two test procedures to test again.

[0049] sample type

[0050] The test results showed that the test results of 16 serum samples were inconsistent. Taking the detection results of the competitive inhibition ELISA detection kit as a reference value, the sensitivity of the double-antigen sandwich ELISA method can reach 99.8%, the specificity can reach 95.7%, and the coincidence rate can reach 98.3% (Table 2).

[0051]

[0052]...

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Abstract

The disclosed dual-sandwich enzyme-linked immunologic diagnosis agent box for hepatitis-B core antibody comprises: a micro-porous reaction plate composed by the hepatitis-B core antigen recombinant with purity more than 90% and 5*104~5*105u / mg titer, and the enzyme compound composed by the hepatitis-B core antigen recombinant marked by HRP. This invention has high specificity and sensitivity and convenient to qualitative or semi-qualitative detection.

Description

technical field [0001] The invention relates to a double-sandwich method ELISA diagnostic kit for detecting hepatitis B core antibody, which is suitable for qualitative or semi-quantitative detection of hepatitis B core antibody. Background technique [0002] Hepatitis B virus (HBV) is the pathogen that causes hepatitis B in humans and is one of the most widespread deadly viruses in the world. According to statistics, more than 3.5 billion people in the world are carriers of chronic hepatitis B virus [Zuckerman J N, Zuckerman A J. Current topics in hepatitis B. J Infect, 2000, 41 (2): 130-136], among which China has 120 million; and about one million people die every year worldwide due to HBV infection [Parkin, D.M, Pisani P, Munoz N and Ferlay J. The global healthburden of infection associated cancers. Cancer Surv, 1999, 33: 5 ~33.]. [0003] Since there is no effective means to treat hepatitis B at present, vaccination and early detection and di...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/547
Inventor 黄健邓丽娟严兵
Owner SHENYANG HUIMIN BIOTECH CO LTD
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