Objective:To study the influence of osteoclast generation and function in different methods of osteoclast culture in vitro. Methods:We used 3 methods of osteoclast culture in vitro:the group A,10 ng/ml M-CSF was added in mice marrow cell for 24 h. Cells which not adhere to the surface were pre-induction with 30 ng/ml M-CSF for 3 d,and then 50 ng/ml M-CSF + 100 ng/ml RANKL were added;the group B,mix culture mice marrow cell with mice osteoblast on 10∶1,and 10-6 mol/L PGE2 and 10-8 mol/L VitD3 were added;the group C,RAW264.7 was added with 100 ng/ml RANKL to inducement. Osteoclastogenesis and their resorption function were examined by TRAP staining and scanning electron microscope (SEM) observation of dentin resorption lacunaes. The gene expressions of NFATc1 and c-Fos were also detected by Real-time PCR. Results:TRAP positive multinuclear cells were observed and resorption lacunaes were formed in each group. However,the group B showed more TRAP positive multinuclear cells and larger amount of resorption lacunaes than the group A and C. The group C showed more TRAP positive multinuclear cells but less amount of resorption lacunaes than the group A. Real-time PCR detection also showed that the gene expressions of NFATc1 and c-Fos were higher in the group B than the group A and C. Conclusion:As the comparison of three methods of osteoclast culture,the group B is better than the group A and C in osteoclastogenesis and resorption function. The group A is better than the group C in the resorption function of osteoclast,but the group C is better than the group A in osteoclastogenesis.