White adipose tissue (WAT) is no longer considered a depot for energy storage in the form of triglycerides, but is a secretory organ that releases factors, known as adipokines, capable of regulating several physiological processes. Alteration of WAT function with subsequent dysfunctional expression and secretion of adipokines plays a key role in the pathogenesis of obesity, diabetes, and other metabolic diseases. For this reason, a deeper understanding of the molecular mechanisms regulating adipocyte function is deemed necessary for planning strategies to treat and prevent obesity and its metabolic complications. This review examines cell culture models currently available for studying adipocyte biology. We focus on advantages, disadvantages and main differences between established preadipocyte cell lines and primary preadipocyte cultures. We revise protocols used to promote adipocyte differentiation and mature adipocytes dedifferentiation into preadipocytes. Finally, we briefly describe co-cultures of adipocytes with other cell types and three-dimensional adipocyte culture systems. These models allow investigation of cell–cell interactions with the cross-talk physiologically occurring between adipocytes and other cell types residing within or outside adipose tissue. J. Cell. Biochem. 110: 564–572, 2010. © 2010 Wiley-Liss, Inc.