Isolation, culture and phenotypical analysis of dendritic cells from human peripheral blood

Objective: This study was designed to establish the methods of isolation, culture and phenotypical analysis of dendritic cells from human peripheral blood. Methods: The procedure involved two steps. The first step was as following. Normal human peripheral blood were isolated by ficoll centrifugation and the interface cells were harvested and cultured in plates. The adherent cells were then cultured with human recombinant granulocyte/macrophage colony- stimulating factor (rhGM-CSF) on plus human recombinant interleukin 4 (rhIL-4) about 7 days. The second step was to put tumor necrosis factor α (TNF-α) in medium. The next day the suspended cells were checked by morphological and FACS. Results: The suspended cells exhibited distinctive morphological features of dendritic cells and expressed HLA-DR、CD1a、CD80、CD83 and CD86 highly, while CD14 dimly on cell surface. Conclusions: The results indicated that by combination of GM-CSF, IL-4 and TNF-α, Large numbers of dendritic cells could be obtained from human peripheral blood. Generation of dendritic cells from human peripheral blood may facilitate further studies on dendritic cell cancer vaccines and their clinical applications.