目的:探讨可用于临床治疗功能成熟的DC体外扩增的优化培养方案.方法:胎牛血清培养基联合细胞因子rhGM-CSF(100ng/mL)和rhIL-4(50 ng/mL)扩增人外周血分离的单个核细胞,细胞培养分别按5× 106/mL、6× 106/mL和 7× 106/mL的密度,加入6孔培养板.第6d加入rhTNF-a(100 ng/mL)联合培养,分别于第6d,第9d和12d收获细胞.从形态学、细胞表面标志方面进行鉴定.结果:显微镜观察,经过9d诱导后,培养细胞具有典型树突细胞外形.流式细胞仪分析,6× 106/mL密度的细胞培养组培养到第9天最宜.结论:细胞具有典型的DC的形态特征,细胞表型及功能实验证实其DC的特性,说明建立的血清培养基联合细胞因子rhGM-CSF、rhIL-4和rhTNF-a体外诱导DC的方法是切实可行的.%Objective: To investigate a culture system for DC expansion. Methods: Using serum medium with rhGM-CSF (l00ng/mL) and rhIL-4 (50ng/mL) for 6 days, rhTNF-a (lOOng/mL) was added on the 6th day of culture. Human peripheral blood were seeding at the concentration of 5x 10VmL, 6×106/mL and 7× 106/mL in 6-well flat-bottomed plates. Suspension cells were harvested at days 6, 9 and 12 and analysed on morphology, phenotype observed. Result: Cells on day 9 of culture displayed DC-like morphology. Phenotype analysis on DCs acquired showed that the cells of 6×106/mL cultured for 9 days contained a higher percentage of DC cells. Conclusion: The results suggested the culture method was feasible.
展开▼
