摘要
牛支原体可引起牛肺炎、乳腺炎和关节炎等多系统炎症，关于其黏附宿主细胞的机理有待深入研究。本试验以牛支原体08M株和牛肾细胞MDBK为研究对象，抗牛支原体多克隆抗体为一抗，FITC标记的兔抗牛IgG为二抗，通过优化牛支原体黏附滴度、黏附时间及一抗、二抗工作浓度，建立牛支原体黏附宿主细胞的间接免疫荧光检测方法。最终确定牛支原体的最适黏附滴度为1×10^9CFU／mL，黏附时间为4h，一抗最适工作浓度为1：250，二抗最适工作浓度为1：250。本试验建立的间接免疫荧光方法可用于牛支原体黏附宿主细胞的检测，为牛支原体体外感染的检测及黏附机理的研究提供有效的技术手段。
Mycoplasma boris （M. boris） causes multisystemic disorder in calves,including pneumo- nia,mastitis and arthritisetc. The mechanism of this pathogen adhesion to host cells is not clear. In the present study,an indirect immunofluorescence assay was developed to detect adherence of M. bovis to host cells. M. boris strain 08M and MDBK cells were used in the assay,anti-Mb IgG as the primary antibody and fluoriescein isothiocyanated conjugate rabbit anti-cow IgG as the secondary antibody. Optimization of reaction conditions was carried out,including the best titer and time of M. bovis adhesion to MDBK cells, the concentration of the primary antibody and the second anti-body. As a result, the best titer and time of M. bovis adhesion to the MDBK cells were 1×10^9 CFU/mL and 4 h,respectively. The optimum working concentrations of anti-M, boris IgG and FITC-labeled rabbit anti-cow IgG were 1 ： 250 and 1 ： 250,respectively. The IFA assay established in this study could be used as a method to detect M. boris adhesion to host cells,and provide an effective technique for the detection of M. bovis in vitro and its adhesion mechanism.
作者
李明霞
陈胜利
郝华芳
赵萍
季文恒
王展慧
刘永生
殷宏
储岳峰
LI Ming-xia;CHEN Sheng-li;HAO Hua-fang;ZHAO Ping;JI Wen-heng;WANG Zhan-hui;LIU Yong-sheng;YIN Hong;CHU Yue-feng(State Key Labotorary of Veterinary Etiological Biology,Lanzhou Veterinary Reasearch Institute,Chinese Academy of Argricultural Science,Lanzhou 730046,Chin)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2018年第7期1310-1314,1345,共6页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31402223,31602088)
国家科技支撑计划资助项目(2015BAD12B02)
国家重点研发计划资助项目(2016YFD0500907)
甘肃省科技支撑计划资助项目(1204NKCA071)
兰州市城关区科技计划资助项目(2012-2-1)
国家肉牛牦牛产业技术体系资助项目(CARS-38)
关键词
牛支原体
黏附
MDBK细胞
间接免疫荧光
Mycoplasma boris
adhesion
MDBK cells
indirect immunofluorescence assay
