Effect of DHG on the in vitro half-life of factor VIIIa. Recombinant factor VIII (20 nM) was activated for 30 seconds with 40 nM thrombin, neutralized with 60 nM hirudin, and immediately diluted 1:2 into the tenase reaction buffer in the absence (•) or presence of 25 (○) or 50 nM (▪) DHG. Residual factor VIIIa activity was determined by sampling into the intrinsic tenase assay as described (see “Materials and methods”) and the data fit to a simple exponential decay. Mean values for factor VIIIa activity were plotted versus time with error bars representing ± SD (n = 3). The k_obs for loss of factor VIIIa activity was 0.16 min^–1 for cofactor alone and 0.18 min^–1 or 0.19 min^–1 for cofactor plus 25 or 50 nM nM DHG, respectively.