The development of adipocytes was studied in primary cultures of rat adipose tissue stromal-vascular cells on collagen-coated glass coverslips. The effects of cell density and serum source on lipoprotein lipase (LPL), esterase and lipid histochemistry were evaluated. With a mixture of fetal calf serum (FCS; 2%), horse serum (2%) and pig serum (PS; 10%), large and loosely arranged clusters of adipocytes developed with time through an increase in cell number and size. An inverse relationship was observed between cell size and the number of adipocytes in a cluster. Lower cell densities were associated with large cells and the densest areas contained smaller cells. Unilocular adipocytes were observed by d 13 after plating and were generally absent from the densest area of the coverslips. Histochemically detectable LPL activity was demonstrable before lipid deposition in adipocyte clusters. A comparison of FCS (10%) and PS (10%) as the only serum sources indicated higher level of adipocyte esterase activity and lipid deposition in PS cultures. Cultures of cells from weanling (21 to 28 d old) and old (18 mo) rats were similar, whereas cells from younger rats (2 to 4 d old) formed denser cultures that contained fewer adipocytes. These adipocytes were small (less than 30 micron) and morphologically homogeneous (all multilocular). When cells from the very young rats (2 to 4 d old) were plated at low densities, an inverse relationship between cell size and number of cells in a cluster was also observed.(ABSTRACT TRUNCATED AT 250 WORDS)