Objective To explore the methodology regarding culture,proliferation and purification of adipose-derived stem cells(ADSCs),and to study their biological characteristics. Methods Fat tissue was obtained from mesentery and subdermal tissues of rats of different lineages.The tissue samples were digested with collagenase to obtain adipose stromal cells.Cells were isolated and purified based on their different reaction to digestion.Immunohistochemistry was used to characterize and identify the harvested cells.Growth of cells was observed with an inverted contrast phase microscope.Results Despite different productivity,adipose stromal cells from different source tissues and different lineages showed identical biological characteristics.After 2-3 days of culture,floating cells became adherent and grew into clusters.After several passages,these cells separated and changed into spindle-shaped.The adherent cells could be further classified into two types.The first type of larger size contained profuse lipid droplets and highly positive by oil red O staining.The second type was of slimmer-shape,with little intracellular lipid and negative oil red O staining.After passages,remaining floating cells,albeit in a small number,continuesly differentiating into cells of the second type,while cells of first type gradually diminished.Assessed by immunohistochemistry,cells of the second type were CD49d( ) and CD106(-),which were characteristic phenotype of adipose stromal cells. Conclusion With modified duration of digestion,our method can be used to culture,proliferate,and purify adipose stromal cells,despite difference regarding productivity existing among different tissue sources and diverse lineages.