中国兽医科学2017，Vol.47Issue(4)：471-475,5.DOI:10.16656/j.issn.1673-4696.2017.04.011
猪肺炎支原体抗原定量ELISA检测方法的建立
Development of a quantitative enzyme-linked immunosorbent assay for the detection of Mycoplasma hyopneumoniae
摘要
Abstract
In order to establish a rapid detection method of the Mycoplasma hyopneumoniae,an double antibody sandwich ELISA method was established using anti-P46 monoclonal antibody as capture antibody and anti-P46 protein polyclonal antibody as detect antibodies.The optimal coating concentration of monoclonal antibody was 2.5 μ g/mL.The optimal dilution of anti-P46 protein polyclonal antibody and HRP-conjugated IgG was 1∶40 000 and 1∶10 000,respectively.Furthermore,the coefficients of variation of inter-assay and intra-assay experiments were below than 10％.The lowest detection quantity of M.hyopneumoniae was 9.754 ng/mL.No cross reactions with Mycoplasma hyorhinis,Mycoplasma hyosynoviae and Mycoplasma flocculare were detected.The established ELISA method was employed to detect artificially diluted samples.The concentration rate of the samples was calculated by directly detecting the samples (single point method) or by detecting a serial of dilutions of the samples(parallel line method),and the latter was more stable.In summary,the established double antibody sandwich ELISA method provides an effective mean for the quantitative detection of Mycoplasma hyopneumoniae,which can be used for antigen detection in Mycoplasma hyopneumoniae inactivated vaccines,and is convenient for quality control of inactivated vaccine production.关键词
猪肺炎支原体/夹心ELISA/抗原定量/P46Key words
Mycoplasma hyopneumoniae/sandwich ELISA/quantitative detection/P46分类
农业科技引用本文复制引用
田瑞雨,熊祺琰,刘茂军,韦艳娜,武昱孜,冯志新,姜平,邵国青..猪肺炎支原体抗原定量ELISA检测方法的建立[J].中国兽医科学,2017,47(4):471-475,5.基金项目
国家自然科学基金面上项目(31370208) （31370208）
江苏省农业科技自主创新资金项目(CX(14)2040) （CX(14）
