人外周血巨噬细胞培养及功能鉴定

摘要

目的：从人外周血单个核细胞（ PBMC）中分离单核细胞，诱导分化成巨噬细胞并鉴定其功能。方法：应用免疫磁珠法从人外周血单个核细胞中分选CD14＋单核细胞，分离后的细胞用流式细胞仪检测其纯度，用含有10％人AB血清和10 ng／ml人M－CSF的IMDM培养基体外培养CD14＋单核细胞，诱导分化成巨噬细胞，并进行形态特征和吞噬功能鉴定。结果：免疫磁珠法能从外周血中分离到高纯度的CD14＋单核细胞，分选前CD14阳性率为10％，分选后CD14阳性率为85．8％。诱导培养7天后巨噬细胞的直径最大可达40～45μm，大部分细胞呈煎蛋状，能有效地吞噬淋巴瘤Raji细胞。结论：从外周血中分离了高纯度的CD14＋单核细胞，诱导形成的巨噬细胞具有吞噬淋巴瘤细胞的功能。%Objective:To isolate monocytes from human peripheral blood mononuclear cells( PBMC) ,induce macrophages,and identify the function of macrophages.Methods:Monocytes were isolated from PBMC using magnetic activated cell sorting( MACS) anti-CD14 microbead.Sorted CD14+and CD14-cells were checked by flow cytometer to evaluate the efficiency of sorting.The sorted CD14+cells were cultured in IMDM media with 10%human AB serum and 10 ng/ml M-CSF for 7 days to generate macrophages,which were identified by morphological features and phagocytosis function.Results:A high purity of monocytes was obtained by MACS anti-CD14 microbead.The percentage of CD14+cells was 10% and 85.8% before and after sorting, respectively.The macrophages were approximately 40-45 μm in maximum diameter and had the fried egg colony morphological features after 7 days culture.The lymphoma ( Raji) cells were efficiently engulfed by macrophages.Conclusion: The high purity of CD14+monocytes is isolated from PBMC and monocyte-derived macrophages efficiently engulfed lymphoma cells.