Dendritic cells (DC) are highly efficient antigen-presenting cells that initiate the primary immune response. Several laboratories have developed culture systems for human DC from peripheral blood monocytes. Most of these studies have used fetal calf serum (FCS) containing culture conditions that are inappropriate for human application. GM-CSF and IL-4 were used to make immature DC. The monocyte-conditioned medium (MCM) was used to induce the final maturation of DC. Using the previously described methods, the quality of MCM has unpredictable variations. Therefore using a defined cocktail of growth factors for the generation of mature DC would be advantageous for experimental as well as clinical purposes. In this study, it is suggested that combinations of both GM-CSF/IL-4 or GM-CSF/IL-13 could be used as the first-step culture to produce immature DC, and that cytokine cocktail (GM-CSF, IL-4, IL-1β, TNF-α, IL-6, PGE₂) was as efficient as MCM for the second step-culture to produce fully maturated DC. Here, we have generated an easily reproducible culture system for DC that allows for the generation of large amounts of immature and mature DC, and we also now have established the method in a FCS-free system that is suitable for clinical use.