副猪嗜血杆菌间接ELISA检测方法的建立  

Establishment and verification of indirect ELISA for detection of antibodies against Haemophilus parasuis

作　　者：刘双红[1] 李大鹏[2] 徐胜奎[2] 谢芳[2] 李刚[2] 李婷[3] 孙斌[1] 王春来[2] 

机构地区：[1]黑龙江八一农垦大学动物科技学院,黑龙江大庆163319 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/动物细菌病研究室,黑龙江哈尔滨150001 [3]东北林业大学野生动物资源学院,黑龙江哈尔滨150040

出　　处：《中国预防兽医学报》2016年第7期576-579,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：副猪嗜血杆菌病和猪传染性胸膜肺炎防治技术研究与示范(201303034)公益性行业(农业)科研专项

摘　　要：为建立一种快速检测副猪嗜血杆菌(HPS)的血清学方法,本实验采用重组表达的HPS细胞致死膨胀毒素B亚基蛋白(r Cdt B)作为包被抗原,并对各反应条件进行优化,建立了检测HPS血清抗体的间接ELISA方法。以该方法检测HPS阳性血清和其他猪病病原阳性血清,除了HPS阳性血清呈阳性外,其余均为阴性,表明该方法特异性良好。批内重复和批间重复的最大变异系数分别小于7%和8%。此外,将Cdt B-ELISA方法与商品化试剂盒Bio Chek HPS-Opp A间接ELISA抗体检测方法对背景明确的116份HPS阳性血清和112份HPS阴性血清进行检测,商品化试剂盒阳性符合率为25%,阴性血清符合率为100%,总符合率为62%,其中Cdt B-ELISA方法对阳性血清的检出率高于商品化试剂盒。应用Cdt B-ELISA方法检测330份临床猪血清样品,结果显示,HPS抗体阳性检出率为11.8%。该方法可以用于HPS的临床检测,为HPS流行病学调查和防控提供了一种快速有效的检测方法。In the present study, an indirect-ELISA （rCdtB-ELISA） for the detection of the antibody against Haemophilus parasuis in serum was established using the prokaryotic expressed subunit of cytolethal distending toxin （rCdtB） as coating antigen. The rCdtB-ELISA was high specificity for detection of the antibody against H.parasuis, but no cross-reactions to the positive sera of TGEV, PRV, PCV2, CSFV, PEDV, PRRSV, E.coli, S.suis2, P.multocida and A.pleuropeumoniae. The coefficient of variation of intra- and inter batch repeatability test was lower than 7% and 8%, respectively. Moreover, comparison of the established rCdtB-ELISA with the commercial ELISA （BioChek HPS-OppA kit） for detection of 116 positive and 112 negative sera to H. parasuis, the results showed that the positive concordance rate was 25%, the negative concordance rate was 100%, and the total concordance rate was 62%. While, a total of 330 clinical sera samples were detected using the rCdtB-ELISA, it showed that the positive rate of H.parasuis was 11.8%.

关 键 词：副猪嗜血杆菌 rCdtB 间接ELISA 

分 类 号：S852.61[农业科学—基础兽医学]