单抗捕捉ELISA检测猪附红细胞体血清抗体方法的建立  

A capture ELISA with McAb for detection of antibodies against Eperythrozoon suis

作　　者：张浩[1,2] 单虎[1] 吴延功[2] 张守富 吴晓东[2] 王志亮[2] 刘佩兰[2] 朱绍辉[1,2] 

机构地区：[1]莱阳农学院动物科技学院,山东青岛266109 [2]中国动物卫生与流行病学中心,山东青岛266032 [3]青岛中仁药业有限公司,山东青岛266329

出　　处：《中国兽医学报》2009年第1期55-58,91,共5页Chinese Journal of Veterinary Science

基　　金：国家“十五”科技攻关资助项目(2004BA514A)

摘　　要：在制备针对猪附红细胞体病的特异性单克隆抗体的基础上,建立了单抗捕捉快速检测猪附红细胞体病抗体的ELISA方法,并对影响ELISA结果的各项反应条件进行了优化。结果表明,猪附红细胞体腹水单抗的最佳稀释度为1∶20 000,猪附红细胞体病抗原工作质量浓度为4.088 mg/L,血清稀释度为1∶40,HRP标记的兔抗猪IgG的工作浓度为1∶5 000。待检血清和酶标二抗的反应时间均为37℃1 h;底物在室温显色时间为10 min。已建立的ELISA方法与PCR平行检测50份样本,总符合率为92%,表明该方法特异性强、快速、敏感性高。用该法检测了各地采集的171份血清,阳性率为40.4%,与镜检的符合率为59.2%;阴性率为47.4%,与镜检的符合率为66.7%,总符合率为67.3%。The procedure of capture ELISA for detection of Eperythrozoon suis antibodies were established with the monoclonal antibodies anginst Eperythrozoon suis,and the best concentrations of the monoclonal antibodies,the Eperythrozoon suis antigens,the serum and HRP-Rabbit anti-swine were 1/20 000,4. 088 mg/L, 1/40 and 1/5 000 respectively. The working time of the serum and HRP-Rabit anti-swine is 37℃ 1 h; the reaction time of TMB is 10 min at room temperature. 50 serum samples were detected paralelly by the capture ELISA and PCR respectively,the accordant rate is 92 %. The capture ELISA is specifically, speedly, sensitively, so it provided a dependable diagnosis method for Eperythrozoon suis. 171 sera samples came from different areas in China was detected by the ELISA kits. The result indicated that the positive rate is 40.4% ,the accordant rate is 59.2% to the detection by optics microscope;the negative rate is 47.4% ,the accordant rate is 66.7% to the detection by optics microscope;the total accordant rate is 67.3%.

关 键 词：猪附红细胞体病 单克隆抗体 单抗捕捉ELISA 抗体检测 

分 类 号：S855.99[农业科学—临床兽医学]