Effect of band 3 phosphorylation on its extractability from whole erythrocytes. Erythrocytes were treated with different o-vanadate concentrations for 30 minutes at 37°C in the presence (lanes 6-10) or absence (lanes 1-5) of Syk inhibitors. Released band 3 was then extracted by incubation in 0.5% Triton X-100 at 0°C for 15 minutes, and the soluble fraction was separated from the insoluble membrane skeletons by centrifugation. Extracted proteins in the supernatant were separated by 8% SDS-PAGE, and the band 3 content of the supernatant was visualized by Western blotting using an anti–band 3 antibody.