重组水痘-带状疱疹病毒gEΔ-Fc融合蛋白在CHO细胞中的表达、鉴定及免疫原性分析

Expression of recombinant varicella-zoster virus gEΔ-Fc in CHO cells and analysis of its immunogenicity

目的应用CHO细胞表达系统表达重组水痘-带状疱疹病毒(varicella-zoster virus,VZV)gEΔ-Fc融合蛋白,为筛选重组带状疱疹疫苗候选抗原提供参考。方法将含有编码gEΔ-Fc基因的真核表达质粒转染CHO细胞,经MSX加压筛选和有限稀释法挑选单克隆,获得分泌表达gEΔ-Fc融合蛋白的CHO细胞株。MabSelect SuRe亲和层析纯化gEΔ-Fc融合蛋白,ELISA法对gEΔ-Fc融合蛋白与Fc受体结合进行鉴定,流式细胞术检测DC2.4细胞对抗原的吞噬效果。gEΔ-Fc融合蛋白免疫BALB/c小鼠后,ELISA法检测小鼠血清抗体效价。结果获得表达gEΔ-Fc融合蛋白的CHO细胞株;流式细胞术证明DC2.4细胞对gEΔ-Fc融合蛋白的吞噬强于gE;gEΔ-Fc融合蛋白可以诱导BALB/c小鼠产生高滴度的抗VZV抗体。结论应用CHO细胞表达的重组VZV gEΔ-Fc融合蛋白具有较好的免疫原性。本研究为筛选重组带状疱疹疫苗候选抗原提供了数据支持。

Objective To express the recombinant varicella-zoster virus(VZV)gEΔ-Fc fusion protein using CHO cell expression system,and provide reference for screening candidate antigens of recombinant herpes zoster vaccines.Methods A eukaryotic expression plasmid containing the gEΔ-Fc gene was transfected into CHO cells.Monoclonal cells were selected by methionine sulfoximine(MSX)pressure screening and limited dilution method to obtain the CHO cells secreting and expressing the gEΔ-Fc fusion protein.The expressed gEΔ-Fc fusion protein was purified by MabSelect SuRe affinity chromatography.The binding activity of gEΔ-Fc fusion protein to Fc receptors was identified by ELISA.Flow cytometry was used to detect the phagocytosis of antigens by DC2.4 cells.Antibody titers in serum samples of BALB/c mice immunized with the gEΔ-Fc fusion protein were detect by ELISA.Results A CHO cell line stably expressing the gEΔ-Fc fusion protein was obtained.Flow cytometry suggested that the phagocytotic activity of DC2.4 cells against the gEΔ-Fc fusion protein was stronger than that against gE.Moreover,the gEΔ-Fc fusion protein could induce BALB/c mice to produce high titers of specific anti-VZV antibodies.Conclusions The recombinant VZV gEΔ-Fc fusion protein expressed in CHO cells had a good immunogenicity.This study provided reference for screening candidate antigens of recombinant herpes zoster vaccines.