Fig. 3. Binding of AP-2 by Syt IX. Twenty microgram of GST, GST-Syt IX-C2A or GST-Syt IX-C2B were immobilized on glutathione-Sepharose beads and incubated in the absence or the presence of Ca²⁺ (3 mM) as indicated, for 4 hours at 4°C with RBL cells lysates (500 µg). Bound proteins were eluted by sample buffer, resolved on SDS-PAGE and analyzed by western blot using anti- adaptin (AP-2) antibodies.