Dendritic cell (DC) is the most powerful inducers and regulators of immune responses, responsible for interaction within the immune system. The ability of DC to induce or regulate/suppress immune responses led to attention in their immunotherapeutic use in various disease types. The aim of the presented study was to generate in vitro CD14+ enriched peripheral blood monocytes (PBMCs) following the culture with human monocyte-derived dendritic cell serum-free differentiation media over ten days with phenotypical analysis and morphological identification for functional studies. In vitro, peripheral blood of human donors samples were collected for the purification of blood CD14+ monocytes that represent the most common origin for DC precursors based on ficoll density gradient separation and human monocyte-derived dendritic cell enriched differentiation medium. After PBMCs isolation, the cell viability, cell yield were determined, the monocyte-derived dendritic cell surface marker expression was detected by flow cytometric analysis after staining with specific fluorescence-conjugated monoclonal antibodies. in vitro culture and differentiation of human monocytes into DCs and their subsequent maturation into mature DC constitute a critical first step for different downstream analysis ranging from an understanding of both human DC biology and function to their use in clinical diagnostics and therapeutic design for different disease.