Objective To compare the rat osteoblast proliferation and differentiation in the mineralization condition and non-mineralization condition, and to prove the feasible time of adding the mineralization liquid to the cell surroundings. Methods SD rat′s calvarial osteoblast was primary cultured and passaged to the 4th generation. After proliferation stabilized, the 4th generation rat′s osteoblast was cultured in the mineralization condition and nonmineralization condition. The cellular modality was observed by inverted microscopy analyzing system. Proliferation was described by MTT chromatometry and growth curve. Expression of alkaline phosphatase（ALP） was proved by alkaline phosphatase staining, mineralized nodus alizarin red staining and enzyme -linked immunosorbent assay （ELISA）. The cell apoptosis and proliferation were examined by flow cytometry. Results The proliferation of osteoblast went to the flat stage about in the 8th day. After in the mineralization condition 3 weeks, a number of mineralized nodus formed. After the osteoblast proliferation almost stabilized, the mineralization group and the nonmineralization group had the similar proliferation index（PI）, but in the mineralization group, the expression of alkaline phosphatase of the osteoblast was more and kept longer than the last group. Conclusion After the osteoblast proliferation almost stabilized, adding the mineralization liquid would not influence the proliferation, but accelerate the osteoblast alkaline phosphatase expression and mineralized nodus amount. It proved that the feasible time to add the mineralization liquid was after the cell proliferation almost stabilized.
