Fig. 1

Differentiation of primary human osteoclasts. A Differentiation scheme of monocytes to osteoclasts. B Monocytes were isolated from peripheral blood and differentiated for 7 days to macrophages in the presence of M-CSF either in polystyrene cell culture plates (standard method) with a medium change every 3 days or in Teflon bags (novel method) with no medium change required. For terminal osteoclast differentiation, cells were either harvested from the bags and seeded at the desired cell density in cell culture plates, or kept adherent in plates. In both approaches, cells were cultured for an additional 7 days in a medium containing both M-CSF and RANKL which was changed every 3 days. C + D, Flow cytometry profile and gating strategy (C) including corresponding quantification (D) of the monocyte preparations. E Expression of surface marker proteins CD14, CD16, CD11b, CD163 and HLA-DRB1 was assessed on monocytes from n > 3 donors. Shown is one representative histogram for each marker. Blue: antibody-of-interest, grey: isotype control