肠出血型大肠埃希菌免疫PCR检测方法的建立

Establishment of Immuno-PCR technique for detection of E.coli O157:H7

目的建立检测肠出血型大肠埃希菌O157:H7的免疫PCR技术,确定该方法的灵敏度和特异性。方法链亲和素桥联生物素化的二抗和双链DNA指示分子,构建桥联系统,进而建立免疫PCR技术,通过PCR扩增指示分子检测O157:H7。结果免疫-PCR技术最少可检测到10 CFU/ml的纯培养菌,灵敏度较ELISA提高103倍,非O157:H7菌株检测结果均为阴性。结论免疫-PCR技术具有较高的灵敏度和特异性,操作简便、快速,可作为肠出血型大肠埃希菌O157:H7的检测方法。

Objective Immuno-PCR was established for detecting E.coli O157：H7,and to determine the sensitivity and specificity of this method.Methods Streptavidin was used to attach biotinylated DNA and biotinylated second antibody.The detection was finished by application of the directed molecular.Results The lowest number of O157：H7 detected by Immuno-PCR was 10CFU/ml bacteria.The sensitivity was about 103 times higher than ELISA.The E.coli O157：H7 strains were positive,while non-E.coli O157：H7 and other bacteria were negative.Conclusions Immuno-PCR could be used for the detection of E.coli O157：H7 because of its high sensitivity and high specificity,simplicity and rapidity.