Inefficient decryption of TF^C209A procoagulant activity. (A) Human umbilical vein endothelial cells were cotransduced with PAR2 adenovirus and the indicated TF virus dose (calculated as particles per cell [ppc] based on protein concentration of the virus preparation). TF cell-surface proteolytic activity was quantified by measuring the rate of FXa generation after addition of 10nM FVIIa and 100nM FX. Subsequently, cells were lysed in 15mM octylglucoside and, after dilution to 5mM octylglucosides, the rate of FXa generation was measured upon addition of 20nM FVIIa and 100nM FX. For Western blotting, cells were directly lysed in nonreducing sample buffer and transfers were probed with polyclonal antibody to human TF. (B) Cells were transduced with the indicated TF virus doses in the presence of 200 ppc empty Ad5 control or PAR2 virus, followed by FXa generation assay with 10nM FVIIa and 100nM FX in octylglucoside lysates, as described above. Transfers were probed with polyclonal antibody to human TF or, as a loading control, to integrin β1.