人冠状病毒SARS-CoV、229E和OC43核衣壳(N)蛋白单克隆抗体的制备及3株人冠状病毒N蛋白的抗原相关性观察

Antigenicity analysis of nucleocapsid proteins of 3 human coronaviruses SARS-CoV,229E and OC43 with their monoclonal antibodies

目的通过制备和鉴定人冠状病毒SARS-CoV、229E和OC43核衣壳(N)蛋白单克隆抗体(mAb),观察3株人冠状病毒的N蛋白抗原相关性。方法分别用基因重组SARS-CoV、229E和OC43N蛋白免疫BALB/c小鼠,制备mAb,并采用ELISA间接法、免疫印迹和免疫荧光进行筛选和鉴定,同时用mAb分析3株人冠状病毒N蛋白之间的免疫交叉反应。结果获得多株抗SARS-CoV、229E和OC43 N蛋白mAb杂交瘤细胞株,ELISA间接法、免疫印迹和免疫荧光结果均显示,所有的mAb仅与各自病毒的N蛋白特异性反应,而在3株人冠状病毒N蛋白抗原之间不产生免疫交叉反应。结论获得的特异性抗人冠状病毒N蛋白mAb为进一步研究3株人冠状病毒的抗原决定簇相关性奠定了基础。

Objective To prepare and characterize monoclonal antibodies （mAbs） against the recombinant nucleocapsid （N） protein of 3 human coronaviruses SARS-CoV, 229E and OCA3 and study the antigenic relationship between the 3 N proteins. Methods BALB/c mice were immunized with the recombinant N proteins of SARS-CoV, 229E and OC43 to obtain the mAbs by means ofhybridoma. Screening and identification of the mAbs were performed using indirect enzyme-linked immunosorbent assay （ELISA）, Western blotting and indirect immunofluorescence assay. Cross-reactivity between the N proteins of the 3 coronaviruses was analyzed with the prepared mAbs. Results The mAbs against the recombinant N proteins of SARS-CoV, 229E and OCA3 were obtained, which reacted specifically with the corresponding viral N protein as shown by indirect ELISA, Western blotting and indirect immunofluorescence assay. No cross-reactivity was found between the 3 N proteins. Conclusion The prepared mAbs against the recombinant N proteins may provide valuable assistance in studying antigenic relationships of N proteins between the 3 human coronaviruses.