Fig. 4

RANKL⁺ effector memory B cells in concert with tumor necrosis factor alpha (TNF-α) facilitate osteoclastogenesis. a–c RAW-Venus1 cells were co-cultured with switched-memory B cells at 2.25 × 10⁴ cells/well for 2 days with or without BCR/CD40 stimulation, interferon gamma (IFN-γ, 20 ng/ml), TNF-α (100 ng/ml) or monoclonal antibody against receptor activator of nuclear factor kappa-B ligand (αRANKL, 100 ng/ml). Venus positive cells were detected using the fluorescein isothiocyanate (FITC) channel. A field equaled an area of 0.39 mm² (magnification × 20). c Venus positive cells co-localized with switched-memory CD19⁺ B cells from HC. d Venus positive cells were counted in three randomly selected fields of three to five independent experiments. Box plots represent the median values, quartiles and extremes (n = 9, *P < 0.05, **P < 0.005). BCR B-cell receptor, DAPI 4’6-diamidino-2-phenylindole, No stim No stimulation, NS not significant