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目的在获得了具有免疫原性的SARS冠状病毒S1蛋白片段的基础上,制备和鉴定特异性抗该段S1蛋白单克隆抗体(mAb)。方法原核表达含S蛋白受体结合区的SARS冠状病毒S1蛋白片段S1c(N端249-667氨基酸残基),其免疫原性经SARS病人恢复期血清鉴定后免疫BALB/c小鼠,按常规方法制备单克隆抗体,并采用ELISA间接法、免疫荧光和免疫印迹进行筛选和鉴定。结果筛选出3株特异性针对SARS冠状病毒S1蛋白N端249-667的mAb杂交瘤细胞株,IgG亚类鉴定1株为IgG1,2株为IgG2a,经免疫荧光鉴定与人冠状病毒株229E和OC43无交叉反应。结论获得3株抗SARS冠状病毒S蛋白受体结合区特异性单克隆抗体,为建立新的SARS冠状病毒检测方法的和进一步研究S蛋白的功能奠定了基础。
OBJECTIVE To prepare and identify a monoclonal antibody (mAb) specific for the S1 protein of S1V on the basis of obtaining immunogenic S1P fragment of SARS coronavirus. Methods S1c fragment of S1S (N terminal 249-667 amino acid residues) of SARS coronavirus containing pro-binding domain of S protein was prokaryotically expressed. BALB / c mice were immunized with the immunogenicity of SARS coronavirus by convalescent sera. Methods Monoclonal antibodies were prepared and characterized by indirect ELISA, immunofluorescence and immunoblotting. Results Three mAb hybridoma cell lines specific for the N-terminal 249-667 of SARS-CoV S1 protein were screened. One IgG subclass was identified as IgG1 and the other two were IgG2a. The results of immunofluorescence staining showed that the McAb hybridized with human coronavirus strain 229E and OC43 no cross-reaction. Conclusion Three monoclonal antibodies against SARS-CoV S protein receptor binding region were obtained, which laid the foundation for the establishment of a new SARS-CoV detection method and further study on the function of S protein.