Characterization of cultured mast cells. (A) Fetal liver-derived mast cells express Apaf-1 and caspase-9, as determined by Western blotting. Lysates of thymocytes and of mast cells cultured from wild-type, Apaf-1^-/-, or caspase-9^-/- fetal liver cells were examined. Probing with an antibody to actin served as a loading control. White lines indicate that lanes of the same exposure of immunoblots have been rearranged for clarity. (B-D) Cells cultured from wild-type, Apaf-1^-/-, and caspase-9^-/- fetal livers in SCF and IL-3 (as described in “Materials and methods”) resemble mast cells in (B) microscopic appearance of cytospins stained with toluidine blue as well as surface expression of (C) FcϵR1a and (D) c-Kit. In panels C and D, solid histograms represent staining with antibodies specific to FcϵR1a (C) or c-Kit (D), respectively, while broken histograms indicate the staining with isotype-matched control antibodies.