Figure 2.

Characterization of GVs, immunoisolated from cultured astrocytes using beads coated with anti-Sb2 monoclonal antibodies. A, Electron micrographs of the bead fraction, showing vesicles bound to the surface of the beads. Ctl, Immunobeads coated with mouse IgG that were processed in parallel; no bound vesicles are detectable. Arrows, Bead-bound vesicles. Scale bars, 100 nm. B, Size distribution of the immunoisolated vesicles. A total of 300 vesicles was measured, with the numbers falling into the respective categories indicated in the histogram. The black curve shows a Gaussian fit of the size distribution. C, Astroglial LSS and bead-bound material (IP) were immunoblotted for Sb2. Equal proportion of samples was loaded showing that a major fraction of Sb2-positive organelles was recovered in the bead fraction. Light and heavy chains correspond to light and heavy chains of mouse control (lane 2) and monoclonal anti-Sb2 antibodies (lane 3). D, E, Immunoblots of the fractions shown in C for various marker proteins specific for SVs (D) and subcellular compartments (E). Sph, Synaptophysin; Stg 1, synaptotagmin 1.