Objective The objective of this study was to isolate, characterize, and differentiate ferret adipose?derived stem cells (ADSCs). Methods Subcutaneous fat tissues from ferret abdomens were obtained via germ?free procedures, mechanically minced into 0. 5 mm3 pieces, and digested with 0. 075% collagenase. Subsequently, ADSCs were cultured in high?glucose DMEM medium containing 10% FBS, antibiotics, and 4 ng/ mL β?FGF. Then, analyses of cell morphology and cell surface antigen expression were made by flow cytometry. Finally, osteogenic, adipogenic, chondrogenic, and neural differentiation were examined using different method. Results We isolated and identified ADSCs from fresh fat tissues of ferrets, which demonstrated a more fibroblast?like morphology. FACS analysis show high expression of CD29 (53. 1%), CD90 (99. 6%) and CD105 (93. 7%), but low expression of CD11b (36. 3%) and CD44 (28. 3%). The ADSCs could be differentiated into adipocytes, osteoblasts, chondrocytes, and neurons by in vitro induction. Conclusions ADSCs can be isolated from the subcutaneous fat tissue of ferrets, and these cells have multi?directional differentiation ability.