Mad2 dissociates from c-Kit after stimulation with SCF plus GM-CSF in MO7e cells. (A) After serum starvation, cells were stimulated with 50 ng/mL SCF plus 2 ng/mL GM-CSF for 3 hours. (B) Cells were unstimulated or stimulated with 50 ng/mL SCF, 2 ng/mL GM-CSF, or SCF plus GM-CSF for 3 hours. Total-cell lysates (TCL) were separated into 2 tubes for immunoblotting and immunoprecipitation, respectively. Cell lysates were immunoprecipitated with control IgG or anti–c-Kit Abs, and then immunoblotted with anti–c-Kit Abs (left panel, top blot) or anti-Mad2 Abs (left panel, bottom blot). To compare the Mad2 proteins, separated cell lysates also were immunoblotted with anti-Mad2 Abs (right panel). The results of 1 representative of 2 experiments are shown. IB indicates immunoblot; IP, immunoprecipitation; −, no cytokine stimulation; S, SCF; G, GM-CSF; and S/G, SCF plus GM-CSF.