fMLP induces normal intracellular signaling in
syk−/− neutrophils. (A) Neutrophils were stimulated with 3 μm fMLP at 1 minute and the change in intracellular Ca2+ concentration followed by a ratiometric fluorescence assay. Where indicated, extracellular Ca2+ ions were chelated by 5 mM EGTA. (B) Neutrophils were stimulated for the indicated times with 3 μM fMLP, lysed, and the phosphorylation of ERK and p38 MAP kinase followed by immunoblotting with the relevant phospho-specific antibodies. Total amounts of ERK and p38 MAP kinase were determined by phosphorylation-independent antibodies. (C) Neutrophils were stimulated with 3 μM fMLP for 30 seconds, fixed, and the intracellular F-actin level determined by staining with fluorescently labeled phalloidin. Phalloidin binding was quantitated by flow cytometry. MFI indicates mean fluorescence intensity.
