Culture and Differentiation of Monocyte Derived Macrophages Using Human Serum: An Optimized Method

Abstract

Background: Monocyte derived macrophages (MDMs) are appropriate in vitro models to study the function of macrophages in immune related diseases. Not only most of the methods in literature for efficient MDM culture and differentiation are expensive but also they require specific equipment. However, there are some reports indicating that monocyte enrichment is possible through attachment. Purification and differentiation of macrophages occurs in media containing human serum or platelet depleted plasma without extra supplementations. Different variables such as incubation time and serum concentration affect this simple MDM preparation method. Objectives: Here we represent an optimized simple method for MDM preparation from peripheral blood mononuclear cells (PBMC). Materials and Methods: To introduce an optimized method we accomplished the present descriptive study. After PBMC isolation and monocyte enrichment in complete RPMI 1640 growth media, macrophages were cultured and differentiated using human serum. Efficient phagocytosis was evaluated using heat-inactivated Escherichia coli followed by SYBR staining. Geimsa staining of macrophages was accomplished to visualize the typical morphology under light microscopy. Results: The derived macrophages have the typical morphology of differentiated macrophages and are able to phagocyte the heat inactivated SYBR stained E. coli. Conclusions: This optimized method is a simple and cost effective method to prepare MDM with typical morphology representations able to phagocytosis efficiently.