人工抗原提呈细胞的制备和应用的研究

目的 制备人工抗原提呈细胞（artificial antigen presenting cell,aAPC）并用它从HLA-A2阳性健康个体外周血单个核细胞（PBMC）中诱导和扩增特异性细胞毒性T淋巴细胞（CTL）.方法 将HLA-A2-EBV四聚体分子和抗CD28抗体分子吸附固定在细胞大小的聚苯乙烯乳胶微球（5μm）表面制成aAPC;采用流式细胞仪表型分析;aAPC和HLA-A2阳性个体人外周血单核细胞进行混合淋巴细胞反应;用HLA-A＊0201-EBV四聚体染色法检测特异性CTL的频率;应用细胞内细胞因子染色法检测特异性CTL功能性细胞因子IFN-γ的分泌;采用LDH释放法检测特异性CTL的特异杀伤活性.结果 流式细胞仪分析显示微球表面吸附有HLA-A2-EBV四聚体分子和抗CD28抗体分子;四聚体检测及细胞内细胞因子染色法检测与经典细胞毒试验结果一致,结果表明aAPC在体外可诱导抗原特异性CTL的生成.结论 aAPC制备成功,并在体外有效地诱导抗原特异性CTL的生成.

The preparation and application of artificial antigen presenting cells

Objective To prepare artificial antigen presenting cells(aAPC)and to induce specific cyto- toxic T cells(CTL)using aAPC from peripheral blood mononuclear cell(PBMC)from HLA-A2~+ normal donors. Methods aAPC was generated by coupling HLA-A2-EBV and anti-CD28 antibody to the surface of the cell-size microbeads.The immunophenotype of aAPC analyzed by FACS.PBMC from HLA-A2~+ donors were co-cultured with aAPCs.Specific CTL response was demonstrated by HLA-A2-EBV tetramer staining,intracellular cytokine stain assay and cytotoxicity assay.Results The analysis of FACS shows that HLA-A2-EBV and anti-CD28 anti- body are immobilized on microbeads.The result of tetramer staining,intracellular cytokine stain assay and cytotox- icity assay show that aAPC successfully induces specific CTL successfully.Conclusion aAPC was prepared and could induce specific CTL in vitro.