Demonstration of the Phe522Cys c-kit mutation and its functional analysis. (A) Direct sequencing of codon 522 of c-kit cDNA synthesized from bone marrow aspirate cells. Arrow denotes the T>G point mutation. Upper panel, patient; lower panel, healthy control. (B) Western blot analysis of phosphorylated Kit protein in Cos-7 cells transfected with wild-type, Phe522Cys, or Asp816Val mutants of Kit incubated with or without SCF and 1 μM imatinib mesylate. (C) In vitro sensitivity of the patient's bone marrow aspirate mast cells to 1 μM imatinib mesylate after 8 days of culture (top row) compared to those of a patient with a codon 816 c-kit mutation (bottom row). The circled populations represent mast cells, with their percentage denoted above the circle.