Effects of PMA on the activation of the tyrosine phosphorylation of the p85 subunit of PI3-kinase induced by GM-CSF in human neutrophils. The cells were simultaneously treated with BSA (0.01%), GM-CSF (4 nmol/L), and/or phorbol esters (90 nmol/L) for 15 minutes at 37°C. Neutrophils were lysed under denaturing conditions and the immunoprecipitation was performed using agarose-conjugated antiphosphotyrosine antibodies. The p85 subunit was revealed by blotting using specific anti-p85 antibodies and quantified by densitometry. Statistical significance was evaluated using unpaired Student's t-test. Mean ± SEM of four independent experiments.