树突状细胞的体外培养及鉴定

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.

CULTIVATED AND IDENTIFIED OF DENDRITIC CELLS IN VITRO

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. ResuIts:The results of immunocytochemical method showed that more than 90% cells expressed CD1a, 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, B7-l.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.