肠出血性大肠杆菌O157∶H7脂多糖抗原的提取鉴定及间接ELISA法的建立

Extraction and identification of E.coli O157∶H7 lipopolysaccharide and development of an indirect ELISA

目的提取并纯化具有强免疫反应性和高特异性的肠出血性大肠杆菌O157∶H7(E.coliO157∶H7)脂多糖(LPS)抗原,并研究其在E.coliO157∶H7 LPS抗体检测中的应用。方法应用热酚水法提取E.coliO157∶H7 LPS抗原,同时收集水相和酚相LPS,并对其产率、纯度及免疫反应性进行分析比较,用酸解法制备O-特异性多糖(O-SP),并以不同的LPS为包被抗原,建立间接ELISA法检测E.coliO157∶H7 LPS抗体。结果大分子量的E.coliO157∶H7 LPS优先溶于酚相,而水相中含有更多的小分子量LPS,与水相LPS相比酚相LPS具有更高的免疫反应性和纯度,酚相LPS经酸解去脂得到的O-SP具有更高的反应特异性。结论酚相LPS和O-SP都可用于E.coliO157∶H7 LPS抗体的ELISA检测,O-SP具有更高的特异性,是检测E.coliO157∶H7 LPS特异性抗体的理想材料。

To obtain lipopolysaccharide（LPS）of Escherichia coli O157： H7（E.coli O157： H7）and study the application of LPS in detection of anti-O157： H7 LPS antibody,in the present study the LPS was extracted from E.coli O157： H7 by the hot phenol-water method and both the aqueous and phenol phases were collected.The yield,purity and immunoreactivity had been investigated,and then the phenol phases LPS was hydrolyzed by acetic acid for getting O-Specific Polysaccharide（O-SP）.Indirect ELISA with different LPS as antigen was performed to detect anti-O157： H7 LPS antibodies.The results showed that the high molecular mass LPS was preferentially extracted by the phenol phase,and the aqueous phase contained more low molecular mass LPS.Compared with aqueous phases LPS,the phenol phases LPS has higher purity and immunoreactivity.O-SP has higher reaction specificity,it could be used to detect antibodies of E.coli O157： H7 LPS.