Figure 3. Osteoclast assay to determine retention of RANKL binding of OPG-mutants.
Primary macrophage cells were cultured in the presence of 200 ng of recombinant OPG, OPG-wt or indicated OPG-mutants in the presence or absence of 60 ng of recombinant RANKL. As a positive control, macrophage cells were cultured with RANKL alone. Tartrate-resistant acid phosphatase (TRAP) staining of macrophage cell cultures were performed 7–14 days after cells were cultured in the presence of OPG-wt or OPG-mutants and RANKL (A). Quantitative analysis based on the number osteoclast seen in 10 random fields (B) (*p<0.005 compared to culture with RANKL only).