新型冠状病毒N蛋白优势表位区段抗原的克隆表达及其诊断价值初步评价

Cloning and Expression of the Predominant Epitope Seg⁃ment Antigen of the New Coronavirus N Protein and Pre⁃liminary Evaluation of its Diagnostic Value

目的:制备高活性新型冠状病毒N蛋白的优势表位区段抗原,并初步评价该抗原在新型冠状病毒肺炎诊断中的应用价值。方法:利用生物学软件分析N蛋白的B细胞表位分布,选取含有优势表位的抗原区段,然后用分子生物学技术进行克隆表达获得纯化抗原,ELISA初步评价该优势表位区段抗原的检测性能。结果:经过分析筛选确定15~215 aa为优势表位抗原区段,并获得了高效表达抗原。经过Biocore结合实验,确定优势表位区段抗原NP15-215可以与新型冠状病毒肺炎患者血清样本发生特异性抗原抗体反应。基于该抗原建立的IgM和IgG间接ELISA方法,对临床样本的检测灵敏度分别为60.87%、95.65%,特异性分别为90%、100%。NP-IgM检测的AUC值为0.797,NPIgG检测的AUC值为0.998。结论:优势表位区段抗原NP15-215可用于NP抗体检测,利用该抗原建立的NP-IgM/IgG间接ELISA检测方法可以很好地区分新型冠状病毒肺炎患者和健康者。

Objective:The dominant epitope fragment antigen of novel coronavirus virus NP antigen was prepared and its application value in the diagnosis of COVID-19 was preliminarily evaluated.Methods:The B cell epitope distributions of NP antigen were analyzed by BIOSUN software,and the antigen fragment containing the dominant epitope was selected.Then the dominant epitope fragment antigen was cloned and expressed,and it's detection per⁃formance was preliminarily evaluated by ELISA.Results:NP antigen fragment of 15~215 aa was selected and highly expressed.The dominant epitope segment antigen NP15-215 can react with specific antibody in serum sam⁃ples of patients with new coronavirus pneumoniaby Biocore binding experiments.The sensitivity and specificity of NP-IgM were 60.87%and 90%,while the sensitivity and specificity of NP-IgG were 95.65%and 100%.The AUC value of NP-IgM was 0.797 and that of NP-IgG was 0.998.Conclusion:The dominant epitope fragment an⁃tigen NP is the preferred antigen used for NP antibody detection.The indirect ELISA detection methods of NPIgM/IgG established basing NP antigen can be used to distinguish COVID-19 patients from healthy controls.