摘要
建立阪崎肠杆菌(Enterobacter Sakazakii,ES)特异性抗体间接ELISA检测方法,以期作为融合后杂交瘤细胞株的筛选体系。应用灭活的阪崎肠杆菌菌体为抗原,筛选最佳反应条件,建立检测该病原菌的酶联免疫吸附检测方法。通过对ELISA反应条件优化,确定菌体抗原包被的最适工作浓度为2×105mL-1,4℃包被过夜;5%脱脂奶粉进行封闭;血清最佳稀释比为1∶2 000;酶标抗体最适稀释度为1∶1 000倍;最后确定37℃显色10 min,检测灵敏度达到103mL-1。该方法的建立为阪崎肠杆菌单克隆抗体制备过程中阳性杂交瘤细胞株的筛选提供有效手段和奠定实验基础。
To develop an indirect ELISA for specific antibody against Enterobacter Sakazakii was investigated.This method can be used to screen hybridoma cell lines after fusion. Enterobacter Sakazakii inactivated was applied as antigen, after the optimal reaction conditions were selected , and an indirect ELISA for detection of Enterobacter Sakazakii was established. The optimization of ELISA reactions were followings： the optimal concentration coated was 2x10s mL^-1, and the best coating condition was 4℃ overnight;the blocking buffer was 5% skim milk powder;posi- tive and negative sera dilutions were 1：2 000; the dilution of conjugate was 1：1 000; the TMB substrate was added and incubated at 37℃for 10 min,the sensitivity of Enterobacter Sakazakii was 103 mL^-1. This study demonstrated that the estabhshed ELISA method will provide effective tool for screening hybridoma cell lines secreting monoclonal antibody against Enterobacter Saleazakii and make a foundation for further research.
出处
《中国乳品工业》
CAS
北大核心
2013年第1期51-54,共4页
China Dairy Industry
基金
贵州省卫生厅优秀医学青年科技人才基金(gzwkj2010-2-002)
