Separation of ubiquitinated Syk using an antiubiquitin antibody. Washed platelets were activated with convulxin (100 ng/mL) for 15 seconds. Cells were lysed with RIPA buffer and treated with FK2 antibody for 1 hour at 4°C, followed by Protein A/G PLUS-agarose for an additional hour. Supernatants were treated with anti-Syk using the same protocol. Aliquots were analyzed by Western blotting probing for Syk. Lane A shows total Syk; lane B, the second immunoprecipitate after the supernatant was treated with anti-Syk; and lane C, the material immunoprecipitated by FK2.