Fig. 6.

Cell surface and cytoplasmic nucleolin are carrying Tn antigen. Living unpermeabilised melanoma Ma-Mel-27 cells (after 5 h of culture in fresh medium) were simultaneously incubated with anti-nucleolin mAb MS-3 (dilution 1:25) and with FITC-labeled VVL (green, dilution 1:100) in 10% NGS, 2% BSA/PBS containing 25 mM NaN₃ before PFA fixation (a). For staining of semi-permeabilised (b) or permeabilised cells (c) Ma-Mel-27 cells (after 5 h of culture in fresh medium) were fixed with 3% PFA or 2% PFA/Triton before simultaneous incubation with mAb MS-3 (dilution 1:100) and with FITC-labeled VVL (green, dilution 1:100) in 1% BSA/PBS (overnight, RT), respectively. The bound anti-nucleolin antibody was revealed by Cy3-labeled goat anti-mouse antibodies (red). Nuclei were counterstained with DAPI (blue). Arrows indicate colocalization (yellow) of VVL with nucleolin present on the cell surface. BF; bright field (color figure online)