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目的体外建立慢性乙型肝炎患者永生化的B淋巴母细胞系(LCL)。方法用EB病毒感染自慢性乙型肝炎患者外周血中分离的单个核细胞(PBMC),加入CpGDNA免疫调节基序以诱导B淋巴细胞增殖,环胞菌素A(CysA)抑制T淋巴细胞的活性。光学显微镜下观察LCL的形态特征,利用流式细胞术分析LCL细胞膜表面分子CD19和CD23的表达水平。结果46例患者PBMC经EBV感染4周后,42例转化成永生化B淋巴母细胞系,成功率为91.3%。转化后的B淋巴母细胞体积增大积聚成团,可进一步分裂增殖并长期传代培养。结论转化后的LCL保持了成熟B淋巴细胞的生物学特性,可作为体外研究HBV特异性免疫应答的刺激细胞和靶细胞。
Objective To establish an immortalized B lymphoblastoid cell line (LCL) in patients with chronic hepatitis B in vitro. Methods Mononuclear cells (PBMCs) isolated from peripheral blood of patients with chronic hepatitis B were infected with Epstein-Barr virus (EBV), and CpGDNA immunomodulatory motifs were added to induce the proliferation of B lymphocytes. Cystatin A (CysA) inhibited the activity of T lymphocytes . The morphology of LCL was observed under optical microscope. The expression of CD19 and CD23 on the surface of LCL were analyzed by flow cytometry. Results After PBMCs were infected by EBV for 46 weeks in 46 patients, 42 cases were transformed into immortalized B lymphoblastoid cell line with a success rate of 91.3%. The transformed B lymphoblasts accumulate in volume and accumulate into clusters, which can be further divided and proliferated and subcultured in long term. CONCLUSION: The transformed LCL retains the biological characteristics of mature B lymphocytes and can be used as a stimulator and target for HBV-specific immune responses in vitro.