目的通过构建登革2型病毒B株E基因区1～476 bp的原核表达载体,进行原核表达,并制备单克隆抗体,为研制登革病毒诊断试剂奠定基础。方法将登革2型病毒B株E基因序列克隆入原核表达载体pET28a(+),构建原核表达重组体pET28a(+)-E,将重组表达载体转化BL21(DE3)菌株进行原核表达。纯化后的蛋白免疫BALB/c小鼠,经融合、筛选制备特异性单克隆抗体。结果成功构建了pET28a(+)-E原核表达重组质粒,SDS-PAGE分析表明,E基因区部分序列获得高效表达,获得了1株持续分泌抗E蛋白抗体的杂交瘤细胞株8B3,该单抗为IgG2a亚类。结论成功制备了抗E蛋白mAb,为对登革病毒引起感染的早期诊断提供了有力的工具。 OBJECTIVE: To construct a prokaryotic expression vector of 1 ~ 476 bp in E gene region of Dengue 2 virus B strain and prokaryotic expression, and to prepare monoclonal antibody, which will lay the foundation for the development of Dengue virus diagnostic reagent. Methods The gene sequence of Dengue 2 virus B was cloned into prokaryotic expression vector pET28a (+). The prokaryotic expression recombinant pET28a (+) - E was constructed and the recombinant expression vector was transformed into E. coli BL21 (DE3) for prokaryotic expression. The purified protein was immunized BALB / c mice, fusion, screening and preparation of specific monoclonal antibodies. Results The prokaryotic expression plasmid pET28a (+) - E was successfully constructed. SDS - PAGE analysis showed that the partial E gene sequence was highly expressed and a hybridoma cell line 8B3 secreting anti - E antibody was obtained. IgG2a subclass. Conclusion The anti-E protein mAb was successfully prepared and provided a powerful tool for early diagnosis of dengue virus-induced infection.