Objective To introduce a detailed protocol for isolating and expanding mesenchymal stem cells(MSC) from Wistar rat compact bones.Methods Wistar\'s MSC were isolated and culture-expanded from bone marrow by Percoll density gradient centrifugation or from compact bone debris by digestion with collagenase typeⅡ.The morphological and proliferation features between the two populations of MSC were compared.The in vitro osteogenesis and adipogenesis were identified by intracellular alkaline phosphatase activity and droplets of lipids respectively.Results MSC were successfully isolated and culture-expanded by two different methods.Both of these two populations took fibroblast-like morphology and exhibited similar proliferative activity.They could be induced into adipocytes and osteoblasts in vitro under the defined conditions,which were revealed by positive Oil-Red O and alkaline phosphatase staining.Conclusion Both bone marrow and compact bones could be served as the sources for isolating rat MSC,and the biological properties of the harvested cells are comparable.