FLT3 activation in cells expressing FLT3-ITD Tyr→Phe mutants. Tyrosine phosphorylation was examined in Ba/F3 (A) or 32D (B) cell lysates that were starved of IL-3 for 4 hours. Cell lysates were immunoprecipitated with an anti-FLT3 antibody and subjected to SDS-PAGE separation. Immunoblot analysis was conducted with an antiphosphotyrosine (4G10) antibody (top rows). The blots were then stripped and reprobed with anti-FLT3 antibody (bottom rows).