The effect of the SHIP constructs on SF-induced PIP₃ levels in SHIP^−/− BMMCs.

(A) PIP₃ levels were determined, as described in “Materials and methods,” following a 2-minute stimulation of SHIP^−/−, SHIP^+/+ and SHIP^−/−BMMCs expressing the WT-SHIP construct with 400 ng/mL SF. (B) PIP₃ levels were determined after 2 minutes of stimulation with 400 ng/mL SF in SHIP^−/− BMMCs expressing the indicated SHIP constructs. Because the PI-4,5-P₂ levels were unaffected by either the presence or absence of SHIP or by SF stimulation, they were used as a loading control for the Partisol 10 SAX high-performance liquid chromatography column runs. To reflect this standardization the results are expressed as PIP₃/PI-4,5-P₂ ratios. Each point is the mean ± SE of a minimum of 4 determinations.