Analysis of integrin αIIbβ₃–mediated platelet adhesion and spreading. Platelets from wild type (WT), PI3Kγ^KD (γKD), and PI3Kβ^KD (βKD) mice were plated on glass coverslips coated with fibrinogen for 10, 30, and 60 minutes. Nonadherent cells were removed, and adherent platelets were fixed, permeabilized, and stained with TRITC-conjugated phalloidin. Both platelet adhesion, evaluated as number of counted adherent platelets, and platelet spreading, as number of adherent cells with lamellipodia extension were quantified by fluorescence microscopy analysis. Representative images at 40× magnification of adherent platelets from the 3 genotypes are reported in panel A. The column on the right reports enlarged images (100× magnification) of adherent platelets after 60 minutes to better evidentiate differences in the morphology of spreaded cells. Quantitative analysis of adhesion of wild type (■), PI3Kγ^KD (□), and PI3Kβ^KD () platelets are reported in panels B and C, respectively. Platelet spreading is expressed as percentage of adherent platelets with lamellipodia. Data are the means ± SD of 5 different experiments, where adherent and spreaded cells in 5 different fields were counted in each experiment (*P < .05 vs wild-type platelets).