SBDS localizes to the nucleolus. (A) Retroviral SBDS transduction restores SBDS expression by immunofluorescence. DF259.F fibroblasts infected with retrovirus containing empty vector (top) or SBDS cDNA (bottom) were fixed and probed with the anti-SBDS antibody followed by a FITC-labeled secondary antibody. The cells were counterstained with DAPI (4′, 6-diamino-2-phenylindole) to visualize the nuclei. SBDS was located throughout the cell but was particularly prominent in the region corresponding to the nucleolus. (B) Endogenous SBDS localizes to the nucleolus. HeLa cells were fixed and probed with both an anti-SBDS polyclonal antibody and an antinucleolin monoclonal antibody. SBDS was detected with an antirabbit FITC-labeled secondary antibody, and nucleolin was detected with an antimouse Texas-Red (TR) secondary antibody. Cells were visualized by standard fluorescence microscopy (top) or by confocal microscopy (bottom). (C) SBDS nucleolar localization is intact in SD101 fibroblasts. WT control fibroblasts (NMF-100) or SD101.F fibroblasts were analyzed by immunofluorescence for SBDS. Low power, × 63 magnification; high power, × 100 magnification. (D) SBDS localizes to the nucleolus in DF269 lymphoblasts and in myeloid cells. The indicated lymphoblast cell lines or the myeloid leukemia HL-60 cell lines were subjected to immunofluorescent staining for SBDS protein and nucleolin. Nuclei were counterstained with DAPI.