ProcedureIsolation of Flag fusion proteins with Anti-Flag tag antibody Affinity Gel1Thoroughly suspend the Anti-Flag Affinity Gel in the vial; in order to make a uniform suspension of the resin Quickly transfer 10μl of the gel suspension (about 5μl of packed gel volume) to a fresh 15ml-tube2Add 06ml TBS(or PBS) Thoroughly suspend the Anti-Flag Affinity Gel Centrifuge the resin at 10000 rpm for 30 seconds at 4℃ (or 5 minutes at 400-500×g) Remove the supernatant carefully Be sure that most of the wash buffer is removed and no resin is discarded This step should be repeated for 3-4 times3Add 500μl of cells extracts to the washed resin 4Agitate all samples gently for 2 hours at 4℃5Centrifuge the resin for 30 seconds at 10000 rpm at 4℃ (or 5 minutes at 400-500×g) Remove the supernatants6Wash the resin with 05ml TBS(or PBS) until the OD280 of the supernatant liquid<005 This step should be repeated for 4-5 times7Add 10μl of 2×sample buffer to the resin Boil the samples for 5 minutes8Load 4μl of each sample on SDS-PAGE