CXCL12 increases and AMD3100 inhibits osteoclast activity. Splenocytes of three collagen type II/complete Freund's adjuvant-immunized mice were isolated and pooled. Cell suspensions were cultured for 6 days on a quartz substrate coated with a calcium phosphate film in the presence of macrophage colony-stimulating factor (M-CSF, 20 ng/ml) and receptor activator of NF-κB ligand (RANKL, 100 ng/ml). At day 6 media were removed, cultures were provided with fresh media and stimulated as indicated (M-CSF, 20 ng/ml; CXCL12, 0.5 ng/ml; AMD3100, 25 μg/ml). Cells were removed from the quartz substrate after 2 days and resorption pits were visualized by light microscopy. The resorbed area was measured by a bioquant image analysis system. Bars represent the mean area resorbed by 1 osteoclast (average ± standard error of the mean), measured as the area of 1 resorption pit. The asterisk represents p < 0.001 compared with the M-CSF condition (Mann-Whitney U-test). Representative pictures of resorption pits are shown for the condition stimulated with (b) M-CSF, (c) M-CSF + CXCL12 and (d) M-CSF + CXCL12 + AMD3100. The data in this figure are representative for two independent experiments.