畜牧兽医学报2017，Vol.48Issue(10)：1932-1938,7.DOI:10.11843/j.issn.0366-6964.2017.10.016
猪鼻支原体血清抗体间接ELISA检测方法的建立
Establishment of an Indirect ELISA Detection Method for Mycoplasma hyorhinis
摘要
Abstract
Mycoplasma hyorhinis (M.hyorhinis) is a commensal inhabitant in swine.It can cause polyserositis,pneumonia,arthritis and pericarditis in pigs.Because of the high cross-reactivity among M.hyorhinis and other swine mycoplasmas,there is no specific serological diagnostic method reported.In this study,an indirect ELISA was established by using the variable lipoprotein (Vlp) as the coating antigen.By optimizing the reaction conditions,the optimal concentration of coating antigen was 0.312 5 μg · mL-1 and the serum dilution was 1 ∶ 100,and serum reaction time was 30 min.The optimal dilution at 1 ∶ 20 000 for the goat anti-pig HRP-IgG was determined,and the reaction time was 30 min.The optimal substrate chromogenic time was 10 min.The cut-off value of the assay was 0.27.In the cross-reactivity experiment,the Vlp-ELISA showed no cross-reactivity with common swine pathogens or other swine mycoplasmas.The coefficients of variation of the inter-assay and intra-assay experiments were both below 10％.To investigate the specificity and sensitivity of this method,the positive and negative sera samples were detected by the Vlp-ELISA.After that,a total of 247 clinical samples and sera from the experimentally-infected pigs were tested by the Vlp-ELISA,and the results showed the same tendency with that of Tween 20 extracted membrane protein-ELISA method.In conclusion,the indirect ELISA established in this study was specificity and stability,which can be widely used in clinical detection.关键词
间接ELISA/猪鼻支原体/血清学诊断/VlpKey words
indirect ELISA/Mycoplasma hyorhinis/serum antibodies/Vlp分类
农业科技引用本文复制引用
王佳,熊祺琰,华利忠,刘蓓蓓,甘源,冯志新,刘茂军,武昱孜,邵国青..猪鼻支原体血清抗体间接ELISA检测方法的建立[J].畜牧兽医学报,2017,48(10):1932-1938,7.基金项目
国家重点研发计划(2016YFD0500702) （2016YFD0500702）
