树突状细胞的体外培养及鉴定

CULTIVATED AND IDENTIFIED OF DENDRITIC CELLS IN VITRO

目的:建立树突状细胞(dendritic cells,DCs)体外培养的方法。方法:从正常人外周血分离获得单核细胞,加入rhGM-CSF1000U／ml、rhIL-4500U／ml体外培养。取培养7天的细胞,采用免疫细胞化学方法和流式细胞仪测定细胞表型,并对其进行鉴定。结果:免疫细胞化学方法显示,90％以上培养7天的细胞表达DCs特异性标志CD1a,而单核细胞特异性标志CD14表达小于5％;FACS分析显示,培养7天的细胞高表达HLA-A、B、C,HLA-DR和协同刺激分子CD40、B7-1。表明加入细胞因子体外培养后,由单核细胞生成了DCs。结论:通过该方法对人外周血单核细胞进行体外扩增,能够得到成熟的DC。

Objective： To establish the cultivated methods of Dendritic Cells （DCs） in vitro. Methods： Monocytes were obtained from normal human peripheral blood and cultivated in vitro with rhGM-CSF 1000U/ ml and rhIL-4500U/ml added. After the cells were cultivated 7 days, the phenotype of the cells was identified with immunocytochemical method and flow cytometry. Results：The results of immunocytochemical method showed that more than 90% cells expressed CDla, the specified signals of DCs; while less than 5% cells expressed CD 14, the specified signals of monocytes. The results of FACs analysis showed that the cells which had been cultivated for seven days highly expressed HLA-A, B, C, HLA-DR and co-stimulus molecules CD40, BT-1.The results indicated that monocytes could be incubated into DCs in vitro by adding cytokines.Conclusions： Ripe DCs can be incubated in vitro from monocytes of human peripheral blood.