Figure 5.

Western analysis of sonicated low-density caveolin-rich fractions and immuno-isolated caveolae. (A) Rat lung tissue homogenates were fractionated on a Percoll gradient to isolate plasma membranes (lane PM), followed by sonication and sucrose gradient centrifugation to isolate a caveolin-rich fraction (lane AC). Subsequently, 25 μg of AC was subjected to immuno-affinity isolation using caveolin-1 antibody conjugated magnetic beads separate caveolin-coated vesicles bound to the beads (B) from noncaveolar unbound material (U) in the supernatant. Western analysis was performed on 5 μg of each fraction using antibodies to the indicated proteins. (B) The level of specific proteins in the bound versus the unbound fraction was quantified, and the ratiowas plotted for caveolin and for each of the indicated G proteins.