Fig. 1. Comparison of Rac1 activatino in A5 cells and ETC12 cells adhered to fibrinogen. (A) A5 cells stably expressing IIbß3 and ETC12 cells stably expressing IIbß3728 were incubated in suspension for 2.5 hours and then aliquots of these cells were adhered to fibrinogen (15 µg/ml) for 15, 30 or 45 minutes as indicated. The amount of Rac1 GTP-loading was compared in cell lysates from A5 and ETC12 cells. The levels of Rac1 were detected by western blot in the GST-PAK pull-down assay (top panel) and the lysate (bottom panel) are shown. The average fold change in Rac1-GTP±s.e.m. from five trials is shown in the bar graph (A5 solid bars, ETC12 open bars). The level of Rac1-GTP in the suspended cells was normalized to 1. Significant differences are indicated (*). (B) Surface levels of IIbß3 and IIbß3728 on A5 and ETC12 cells were analyzed by flow cytometry. The x-axis is the FITC fluorescence intensity of A5 or ETC12 cells stained with the directly conjugated P2-FITC monoclonal antibody that recognizes the llb subunit (solid line) or with an IgG-FITC control (dotted line). The y-axis is cell counts. In control samples, human fibroblasts were stained with P2-FITC or IgG-FITC.