Distribution of erythrocyte membrane proteins in microvesicles and lipid rafts.

(A) Erythrocyte membranes (lane 1) and microvesicles (lane 2) were prepared and AChE activity determined as described.1-5Aliquots were loaded on an 11% polyacrylamide gel, with a 4-fold AChE activity in the microvesicular sample, and analyzed by silver staining (upper panel) and Western blotting, as indicated and previously described1-5 (anti–AQP-1 was from Calbiochem, La Jolla, CA). AChE activity is given in arbitrary units. Note the differential distribution of stomatin and flotillin-2. (B) Erythrocyte lipid rafts were prepared by method B1-1 and AChE activity was determined. Aliquots with equal AChE activity of lipid rafts (lane 1) and erythrocyte membranes (lane 2) were loaded on an 11% polyacrylamide gel and analyzed by silver staining (upper panel) and Western blotting, as indicated. CA indicates carbonic anhydrase; GPC, glycophorin C; and AQP-1, aquaporin-1.