抗SARS冠状病毒S1蛋白N端249至667的单克隆抗体的制备与鉴定

南方医科大学学报 ›› 2004, Vol. 24 ›› Issue (01): 1-6.

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抗SARS冠状病毒S1蛋白N端249至667的单克隆抗体的制备与鉴定

温坤¹, 梅亚波¹, 丘立文¹, 廖志勇¹, 袁国勇², 车小燕¹

1. 第一军医大学珠江医院中心实验室, 广东, 广州, 510282;
2. 香港大学微生物学系, 香港

出版日期:2004-01-20 发布日期:2004-01-20
基金资助:
收稿日期:2003-9-15。
基金项目:supported as the Key Medical Research Project for SARS Prevention sponsored by the Ministry of Science and Technology of China and by Guangdong Province

Preparation and characterization of monoclonal antibodies against S1domain at N-terminal residues 249 to 667 of SARS-associated coron-avirus S1 protein

WEN Kun¹, MEI Ya-bo¹, QIU Li-wen¹, LIAO Zhi-yong¹, Yuen Kwok-yung², CHE Xiao-yan¹

1. 第一军医大学珠江医院中心实验室, 广东, 广州, 510282;
2. 香港大学微生物学系, 香港

Online:2004-01-20 Published:2004-01-20

摘要/Abstract

摘要： 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。

Abstract: Objective To prepare and characterize monoclonal antibodies (mAbs) against S1 protein of severe acute respirato-ry syndrome (SARS)-associated coronavirus (SARS-CoV).Methods 6-His-tagged recombinant fragment at N-terminal residues 249 to 667 of SARS-CoV S1 protein including S-protein receptor-binding domain was e xpressed in E.coli. The im-munogenicity of this S1 domain was identified and use d to immunize BALB/c mice for the production of hybridomas. The identification of the mAbs against this S1 domain was performed using indirect enzyme-linked im munosorbent assay (ELISA), indirect immunofluorescence assay (IFA) and Western blotting, respectively.Results Three hybridomas producing mAbs spe-cific to the S1 domain was obtained, with a relative molecular mass of 48 500. None of the 3 mAbs were reactive with human coronaviruses 229E and OC43. Two of the mAbs wer e IgG2a isotype, and the other was IgG1.Conclusion This is the first re-port of mAbs produced against S-protein receptor-binding domain of SARS-CoV. The 3 S1-s pecific mAbs may be useful for further study of the function of the S protein a nd for diagnosis of SARS-CoV infection.

温坤¹, 梅亚波¹, 丘立文¹, 廖志勇¹, 袁国勇², 车小燕¹. 抗SARS冠状病毒S1蛋白N端249至667的单克隆抗体的制备与鉴定[J]. 南方医科大学学报, 2004, 24(01): 1-6.

WEN Kun¹, MEI Ya-bo¹, QIU Li-wen¹, LIAO Zhi-yong¹, Yuen Kwok-yung², CHE Xiao-yan¹. Preparation and characterization of monoclonal antibodies against S1domain at N-terminal residues 249 to 667 of SARS-associated coron-avirus S1 protein[J]. Journal of Southern Medical University, 2004, 24(01): 1-6.

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抗SARS冠状病毒S1蛋白N端249至667的单克隆抗体的制备与鉴定

Preparation and characterization of monoclonal antibodies against S1domain at N-terminal residues 249 to 667 of SARS-associated coron-avirus S1 protein

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