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目的 :探讨定向诱导及纯化树突状细胞 (DC)的方法 ,为进一步研究树突状细胞的功能、特性及临床应用提供技术方法。方法 :利用免疫磁珠法 (MACS)分离纯化脐血CD34 + 细胞 ,在液体培养体系中加入FL、GM CSF、TNF α、IL 4及SCF ,培养 12d后光镜检测细胞形态及流式细胞仪分析表面标志 (CDla、HLA DR及CD80 ) ;利用抗树突状细胞单克隆抗体 (X 11)及免疫磁珠分离纯化树突状细胞 ,流式细胞仪检测纯化后DC的纯度。结果 :经体外定向诱导扩增 ,可成功地将CD34 + 细胞定向诱导为树突状细胞 ,CDla+ 细胞的比例可升至 (2 7.18± 1.5 6 ) %〔对照为 (0 .6 5± 0 .38) % )〕 ,诱导出的DC具有典型的树枝状突起 ,有较高的CDla、HLA DR及CD80表达 ,经免疫磁珠分选 ,树突状细胞 (CDla+ )的纯度可达 (94.6 1± 2 .2 4) %以上。结论 :经特定细胞因子的组合 ,可将CD34 + 细胞定向诱导成DC ,并可通过免疫磁珠法分选出纯度达 90 %以上的DC。
Objective: To explore the method of directed induction and purification of dendritic cells (DCs), and to provide technical methods for further study on the function, characteristics and clinical application of dendritic cells. Methods: The cord blood CD34 + cells were isolated and purified by immunomagnetic beads method. FL, GM CSF, TNFα, IL 4 and SCF were added into the liquid culture system. After cultured for 12 days, the cell morphology and flow cytometry The surface markers (CDla, HLA DR and CD80) were analyzed. The dendritic cells were isolated and purified by anti-dendritic cell monoclonal antibody (X 11) and immunomagnetic beads. The purity of purified DCs was determined by flow cytometry. Results: CD34 + cells were successfully induced to dendritic cells by inducing amplification in vitro. The proportion of CDla + cells increased to (2 7.18 ± 1.56)% [control (0.56 ± 0. 38)%)]. The induced DCs showed typical dendrites with high expression of CDla, HLA DR and CD80. The purity of dendritic cells (CDla +) was up to 94.61 ± 2 .2 4)% above. CONCLUSION: CD34 + cells can be induced into DCs through the combination of specific cytokines, and DCs with more than 90% purity can be sorted by immunomagnetic beads method.