Mo Hsp70 protein based indirect ELISA (enzyme-linked immunosorbent assay) kit and using method thereof

A kit and indirect technology, applied in the field of indirect ELISA kits, can solve the problems of time-consuming and laborious Mo isolation and culture, slow genome research, lack of vaccines, etc., and achieve the effect of high specificity, good effect, and solving the difficulty of cultivation.

Inactive Publication Date: 2015-05-06
GUIZHOU UNIV
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  • Abstract
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  • Claims
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AI Technical Summary

Problems solved by technology

[0003] The prevention and control of Mycoplasma pneumoniae in sheep currently mainly relies on drug prevention, and there is still a lack of reliable vaccines. The reason is related to the time-consuming and laborious isolation and cultivation of Mo and the slow...

Method used

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  • Mo Hsp70 protein based indirect ELISA (enzyme-linked immunosorbent assay) kit and using method thereof

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Embodiment Construction

[0024] Embodiments of the invention:

[0025] The raw materials and formula of the indirect ELISA kit based on Mo Hsp70 protein are:

[0026] Mo standard positive and standard negative sera; the recombinant protein is Mo Hsp70 gene Hsp70 protein was obtained through Escherichia coli Rosseta (DE3) expression system, and the recombinant protein was purified by nickel column method, and the purified target protein was obtained after SDS-PAGE analysis. Diluted by buffer to 1.0μg / 100μL; Coating buffer: Na 2 CO 3 1.59g, NaHCO 3 Dissolve 2.93g in deionized water, dilute to 1000mL, adjust to pH 9.5~9.8; enzyme-labeled secondary antibody: rabbit anti-goat IgG-HRP; 1×PBST buffer: NaCl 8.0g, KCl 0.2g, KH 2 PO 4 0.2g, Na 2 HPO 4 12H 2 Dissolve O 2.9g, Tween-20 0.5mL in 800mL distilled water, adjust pH to 7.2-7.6, and dilute to 1000mL; blocking buffer: 0.3g-0.7g skim milk powder dissolved in 10mL PBST buffer; substrate solution A: Na 2 HPO 4 14.60g, 9.33g citric acid, 0.52g ur...

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Abstract

The invention discloses a Mo Hsp70 protein based indirect ELISA (enzyme-linked immunosorbent assay) kit and a using method of the Mo Hsp70 protein based indirect ELISA kit. The Mo Hsp70 protein based indirect ELISA kit is prepared from the following raw materials: a Mo Hsp70 protein coated ELISA plate, Mo standard positive serum, Mo standard negative serum, an enzyme-labeled secondary antibody, 50-time PBST buffer solution, block buffer solution, substrate solution A, substrate solution B and stop buffer. The Mo Hsp70 protein based indirect ELISA kit can detect the Mo Hsp70 protein antibody of goat serum, can provide key technology for the early monitoring of interstitial pneumonia of sheep and goats, caused by Mo, the immune effect evaluation of vaccines and correlational research, and has important significance on early detection and early control of epidemic diseases caused by the pathogen.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a Mo Hsp70 protein-based indirect ELISA kit and a use method. Background technique [0002] Mycoplasma ovis pneumoniae ( Mycoplasma ovipneumoniae, Mo) is a pathogen that causes interstitial pneumonia (atypical pneumonia) in goats and sheep, which can cause cough, runny nose, progressive emaciation and proliferative interstitial pneumonia symptoms in goats and sheep (Zhang Shuangxiang et al., 2013; Ran Shuangcun, 2010 ; Han Xiao, 2013). In 1963, Mo was first isolated from the lung tissue of Scotland-onset sheep disease by Mackay et al. (Mackay et al., 1963), Spain, Australia, New Zealand, Hungary, Iceland, the United Kingdom, and many countries in Africa have successively reported that Mo causes sheep and goat diseases. occurrence (Xu Jian et al., 2012). Since the first discovery of the pathogen in breeding sheep from New Zealand in 1978 in my country (Hu Jingshao et al., 1982), re...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 程振涛王慧王开功周碧君文明岳筠
Owner GUIZHOU UNIV
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