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目的制备抗出血性大肠杆菌Ο157∶H7(E.coliΟ157∶H7)特异性单克隆抗体(MAbs)。方法福尔马林灭活的E.coliΟ157∶H7免疫BALB/c小鼠,利用细胞融合技术建立分泌抗E.coliΟ157∶H7MAbs的杂交瘤细胞株,对配对较好的6株MAbs用ELISA法测定其免疫球蛋白类及亚类,用ELISA法、凝集法和Westernblot鉴定MAbs的特异性。结果6株MAbs免疫球蛋白均为小鼠IgM。这些MAbs均能与27个E.coliΟ157∶H7菌株发生凝集反应,与部分弗劳地杆菌发生凝集反应,与11株鼠伤寒沙门氏菌、7株伤寒杆菌、2株痢疾杆菌、致病性大肠杆菌、产毒性大肠杆菌、侵袭性大肠杆菌、出血性大肠杆菌Ο26∶H11和Ο111、肠集聚性大肠杆菌、42株非定血清型大肠杆菌、霍乱弧菌Ο1群和Ο139群不发生凝集反应;ELISA结果显示6株MAbs与粪链球菌、变形杆菌、粘质沙雷氏菌、肺炎克雷伯杆菌均无交叉反应;ELISA和Westernblot结果显示,3株MAbs针对E.coliΟ157∶H77酚相脂多糖。结论6株MAbs具有较高的特异性,有可能用于制备检测E.coliΟ157∶H7的病原检测试剂。
Objective To prepare anti-hemorrhagic Escherichia coli O157: H7 (E.coli O157: H7) specific monoclonal antibody (MAbs). Methods BALB / c mice were immunized with formalin-inactivated E.coli O157: H7. The hybridoma cell lines secreting anti-E.coli O157: H7MAbs were established by cell fusion technique. The 6 McAbs with good pairing were determined by ELISA Its immunoglobulins and subclasses were identified by ELISA, agglutination and Western blot. Results All six MAbs immunoglobulins were mouse IgM. These MAbs were able to agglutinate with 27 strains of E. coli O157: H7, agglutinated with part of Bordetella and 11 strains of Salmonella typhimurium, 7 strains of Salmonella typhi, two strains of Shigella dysenteriae, pathogenic Escherichia coli, Enterotoxigenic Escherichia coli, invasive Escherichia coli, hemorrhagic Escherichia coli Ο26: H11 and Ο111, enterotoxigenic Escherichia coli, 42 non-serotype Escherichia coli, Vibrio cholera Ο1 group and Ο139 group no agglutination; ELISA results Six strains of MAbs showed no cross reaction with Streptococcus faecalis, Proteus marcescens and Klebsiella pneumoniae. ELISA and Western blot showed that three strains of MAbs were targeted to E.coli O157:H77 phenolic lipopolysaccharide. Conclusion The 6 strains of MAbs have high specificity and may be used for the preparation of pathogen detection reagents for detection of E. coli O157: H7.