Anatomically separate fat depots differ in size, function, and contribution to pathological states such as the metabolic syndrome. We isolated pre-adipocytes from different adipose depots, omental, subcutaneous and intramuscular, of beef cattle, and cultured in vitro to determine the basis for the variations and attribute these variations to the inherent properties of adipocyte progenitors. The proliferating cells from all depots before the confluence were harvested and the proteome was analyzed by a functional proteomic approach, involving 2-DE and MALDI-TOF/TOF. More than 252 protein spots were identified, selected and analyzed by Image Master (ver 7.0) and MALDI-TOF/TOF. Further, our analysis showed that there were specific differences in proteome expression patterns among proliferating precursor cells from the three depots. Sixteen proteins were found to be differentially expressed and these were identified as proteins involved in cellular processes, heat shock/chaperones, redox proteins, cytoskeletal proteins and metabolic enzymes. The results also enabled us to understand the basic roles of these proteins in different inherent properties exhibited by adipose tissue depots.