Objective To study the effect of fluid flow shear stress on seeded cells for bone tissue engineering. Methods In this study we established a flow chamber perfusion system. The mesenchymal stem cells(MSCs) were implanted into the acellular bone exteracellular matrix, then cultured in the perfusion system under 1.0dyn/cm2 flow shear stress for 7days. Results Alkaline phosphatase(ALP) activity and osteocalcin content, which were two crucial osteoblast genes marker were increased. Cell numbers had no different with the static group. Conclusion It demonstrated that this system is useful for osteoblastic differentiation of bone seeded cells, and this 3D cultured model utilizing fluid flow may be helpful in the culture process of engineering bone.