Generation of an Optimized Dendritic Cell Culture System and Discrimination of the Heterogeneity of the Cultured Cells

Dendritic cell (DC) culture offers a cornerstone for immunological studies; however, its detailed procedures are not very well standardized. This situation has resulted in puzzling ambiguities in attempting to understand the findings resulting from the usage of cultured DCs. This work is designed to optimize the procedure for DC generation, including the initial seeding density of bone marrow-derived mononuclear cells, the application of IL-4 and lipopolysaccharide (LPS), and the methods of cultural medium changing and suspended cell discarding. Furthermore, we discriminate the cultured DCs as subsets of non-adherent, adherent and mixed cells to evaluate their immunological characteristics. The culture system has been proven promising by the DC yield, morphology, purity, viability, and maturation. Remarkably, the cultured DCs’ heterogenic features are identified by cytokine secretion, maturation status and capability to activate allogeneic T cells according to the different subsets; this should be particularly emphasized in immunological investigations when using cultured DCs.