摘要
目的制备大肠杆菌O157∶H7多克隆抗体,比较3种纯化方法对抗体性能的影响,为食品检验、临床和血清学检验O157∶H7奠定基础。方法应用传统方法制备大肠杆菌O157∶H7多克隆抗体,利用盐析法、亲和层析法和抗原吸附法对其进行纯化,采用SDS-PAGE、Bradford法、ELISA等实验技术检测纯化后抗体的纯度、比效价、交叉反应性,从而比较3种纯化方法的纯化效果。结果获得效价1∶131072的大肠杆菌O157∶H7抗血清。亲和层析法所得纯化产物纯度较高,抗体解链为分子量约55 kDa和25 kDa的蛋白,抗原吸附法所得产物纯度次之,盐析法所得产物的纯度较差,有多条杂蛋白条带。抗原吸附法所得纯化产物比效价较高为349.3,并且与其他5株细菌的交叉反应情况相对反应较小。结论亲和层析法和抗原吸附法纯化抗体均有一定的优越性,抗原吸法得到的抗体使用性能较好。
Objective To prepare polyclonal antibody against Escheriehia eoli O157：H7 and compare the effects of 3 different purification methods used in this experiment for the antibody, so as to provide the basis for foods, clinical, and serological assay. Methods After polyclonal antibody against Escherichia coli O157：H7 was made, three different purification methods： salt fractionation, affinity chromatography, and antigen adsorptive process were used to purify the antibody. The purity, specific-activity, and cross-reaction were detemfined by SDS-PAGE, Bradford method, and ELISA. Results The titer of antisera against Escherichia coli O157 ： H7 was 1 ： 131072. The purity of the product obtained by affinity chromatography was the best. The product of affinity chromatography was unwound to protein strands with molecular weight about 55 kDa, and 25 kDa. The purity of the produuct by antigen adsorptive process was the next and it had higher specific-activity up to 349.3 and less cross-reaction. The purity of the product by salt fractionation was poor and had several mixed protein strips. Conclusion Antigen adsorptive process and affinity chromatography both have their advantages to some extent. The performance of the purified product by antigen adsorptive process is better than the latter.
出处
《解放军预防医学杂志》
CAS
北大核心
2008年第4期238-241,共4页
Journal of Preventive Medicine of Chinese People's Liberation Army
基金
国家"八六三"计划资助项目课题(No.2006AA06Z408)
国家科技支撑项目课题(No.2006BAD01B06)
