Thrombin inhibition without or with other plasma inhibitors. (A) Thrombin (4 nM) was incubated either with the indicated concentration of AT (expressed as a percentage of the plasma concentration of 3500 nM) or with the indicated concentration of AT plus plasma concentrations of A1PI, HCII, and A2M. At time points residual thrombin was measured by addition of a thrombin substrate. The rate of thrombin inhibition was calculated from a decay curve fit to the data. As expected, the thrombin-A2M complex retains some ability to cleave substrate. The shaded area indicates the 95% confidence interval of the fit to the data. (B) Substrate cleavage during thrombin inhibition was modeled as a series of ordinary differential equations (equation 3) using the rate constants determined in Figure 1. The model showed good correlation with the experimental data. (C) The model of substrate cleavage includes the contribution of thrombin bound to A2M. Because the IIa-A2M complex is functionally inactive in coagulation, the contribution of IIa-A2M was removed to give a model of residual free thrombin as a function of time (ie, thrombin not bound to an inhibitor). As expected, in the model with only AT, there was no effect of IIa-A2M.