HBsAg真核表达重组质粒在EB病毒转化B淋巴母细胞中的表达

Expression of HBsAg with four eukaryotic expression plasmids in immortalized B-cell line from a chronic hepatitis B patient

目的 探讨HBsAg真核表达重组质粒在乙型肝炎患者EB病毒转化B淋巴母细胞中的表达，建立与该患者相同HLA的HBsAg高效表达细胞系，为检测乙型肝炎患者HBsAg特异性CTL反应提供靶细胞。 方法 取乙型肝炎患者外周血分离的单个核细胞，经EB病毒转化成永生化的B淋巴母细胞系后，用HBsAg真核表达重组质粒pCI-S、pcDNA3.1-S、pMEP4-S、pLXSN-S分别转染B淋巴母细胞，经Hygromycin B或G418筛选，用ELISA检测抗性细胞培养上清液和细胞裂解液中HBsAg。 结果 4种HBsAg重组质粒转染的EB病毒转化细胞，在转录水平上，可检出乙型肝炎表面抗原mRNA的表达；在蛋白水平上，细胞培养上清液和细胞裂解液均可检出HBsAg。 结论 4种HBsAg真核表达重组质粒可在乙型肝炎患者EB病毒转化B淋巴母细胞中稳定表达，为检测该患者HBsAg特异性CTL反应提供相同HLA的HBsAg高效表达靶细胞系。

Objective To establish efficient eukaryotic expression system of HBsAg in immortalized B-cell line from a chronic hepatitis B patient, as autologous HLA target cells for detecting HBsAg-specific cytotoxic T-lymphocyte (CTL) response. Methods Four eukaryotic expression plasmids inserted HBsAg gene were transfected into immortalized B-cell line from a chronic hepatitis B patient, then cells were selected with G418 or hygromycin B. HBsAg in culture supernatants and cell lysates were detected with enzyme-linked immunosorbent assay (ELISA). Results All eukaryotic expression vectors-transfected immortalized B-cells produced HBsAg, which was readily detectable in culture supernatants and cell lysates. Conclusions Transfection of EBV-immortalized B-cell line with four eukaryotic expression plasmids leads to stable expression of HBsAg, which can be used as autologous HLA target cells for detecting HBsAg-specific CTL response. [