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目的:对2019新型冠状病毒(2019-novel Coronavirus，2019-nCoV)核衣壳蛋白(nucleocapsid protein，NP)进行原核表达、纯化与鉴定，并应用于2019-nCoV血清学诊断。方法:合成2019-nCoV NP基因，克隆至pET28a载体中，构建表达质粒，并进行诱导纯化。纯化蛋白经SDS-PAGE、间接ELISA、Western blot(WB)、免疫层析法进行鉴定。优化间接ELISA反应条件，进行血清抗体检测。结果:重组NP经镍柱纯化后SDS-PAGE电泳显示相对分子质量约50×10n 3，与预期一致；间接ELISA、WB表明其能与2019-nCoV感染患者血清特异性结合；用免疫层析法发现对NP检测限为0.2 ng/ml；间接ELISA检测32份2019-nCoV感染者血清及对照血清，发现二者结果可见明显分群。n 结论:原核表达的NP具有良好的免疫原性，可用于制备血清学诊断试剂。“,”Objective:To realize prokaryotic expression, purification and identification of 2019-novel Coronavirus (2019-nCoV) nucleocapsid protein (NP), and apply it to the serological diagnosis.Methods:The synthetic 2019-nCoV NP gene was cloned into the prokaryotic expression vector pET28a to construct expression plasmid, and then purified by Ni-chelating affinity. SDS-polyacrylamide gel electrophoresis (SDS-PAGE), indirect enzyme-linked immunosorbent assay (ELISA), Western blot (WB), and immunochromatography were used to test the purified protein. Indirect ELISA reaction conditions were optimized for serum antibody detection.Results:The relative molecular mass of recombinant NP was about 50×10n 3 after SDS-PAGE electrophoresis, which was consistent with the expectation. Indirect ELISA and WB results showed that it could specifically bind to the serum of patients infected with 2019-nCoV. The detection limit of NP was 0.2 ng/ml by immunochromatography. The sera from 32 patients infected with 2019-nCoV and the control sera were detected by indirect ELISA, and the results showed that they were clearly clustered.n Conclusions:Prokaryotic expression of 2019-nCoV NP has good immunogenicity and can be used for the development of serological diagnostic reagents.