抗原特异性T细胞的体外扩增及其表型和效应功能检测

Proliferation of antigen specific T cells in vitro and analysis of phenotype and function

目的探索一种HLA非依赖性的抗原特异性CD4^＋和CD8^＋T细胞混合型扩增方法。方法以巨细胞病毒（CMV）IE-1抗原为例，IE-1多肽库体外刺激外周血单个核细胞（PBMC）6h后，磁珠筛选IFN-γ阳性细胞，加入放射灭活的PBMC于含IL-2的培养基中培养4周。结果得到I．5×10^9个细胞，扩增效率达2500倍，其中CD8^＋T细胞占92．99％，CD4^＋T细胞6．88％，受阳性靶细胞刺激时，49．2％的T细胞分泌IFN-γ，细胞杀伤率达58％（20：1）。结论该方法可在体外有效扩增T细胞，并从细胞因子的分泌及细胞毒杀伤活性分析，初步认为扩增的T细胞保持其表型和抗原特异性效应功能。

Objective To explore a new peptide-based approach independent of HLA to generate antigen-specific CD4^＋ CD8^＋ T cells. Methods Peripheral blood mononuclear cells（PBMC） were stimulated for 6 h with IE-1 peptide pool. Then the activated IFN-γ secreting cells were tested by immunomagnetic selection. And the selected cells were cultured with radio-inactivated PBMC in medium with 100 IU/ml IL-2 for 4 weeks. Results The generated T cell lines consisted of IE-1 specific CD4^＋ T （6.88%） and CD8^＋ T cells 92.99%, which demonstrated antigen-specific killing and cytokine secretion. Conclusion T cells can be proliferated with this new procedure, and maintain its phenotype and antigen-specific function.