Tyrosine phosphorylated Cbl preferentially associates with CrkII. DA-3-EPO-R cells were depleted of cytokine for 8 hours and stimulated with no factor (lanes 1, 4, 7, and 10), 50 U/mL of murine IL-3 (lanes 1, 5, 8, and 11) or 50 U/mL of human EPO (lanes 3, 6, 9, and 12) for 15 minutes. Following cell lysis, an immunoprecipitation was performed with either an anti-Cbl antibody (lanes 1-3), an anti-CrkII antibody (lanes 4-6), and anti-CrkII antibody followed by an anti-Crk antibody (lanes 7-9), or an anti-Crk antibody (lanes 10-12). The anti-Crk antibody recognizes both CrkI and CrkII. Immune complexes were resolved by SDS-PAGE and blotted to nitrocellulose. The immunoblot was probed with 4G10 monoclonal anti-phosphotyrosine antibody and then stripped and reprobed with either a Cbl polyclonal antibody or a Crk monoclonal antibody. Molecular mass standards are indicated.