Ecarin chromogenic assay--a new method for quantitative determination of direct thrombin inhibitors like hirudin.

A new sensitive and precise method for quantitative determination of direct thrombin inhibitors is described, the ecarin chromogenic assay (ECA). Ecarin is used as the specific prothrombin-activating principle. The cleavage of a chromogenic substrate by meizothrombin is inhibited in a concentration-dependent fashion by direct thrombin inhibitors. For the ECA, the linear measuring range is about 0.1-3.0 microg hirudin/ml plasma. Coefficients of variations between 2.3 and 4% over the whole concentration range were achieved. The ECA has proved to be more sensitive than the compared tests (ecarin clotting time and a thrombin-based chromogenic assay); a detection limit of 0.011 microg hirudin/ml and a quantitation limit of 0.032 microg hirudin/ml were calculated. The ECA is independent of the variations of the coagulation variables fibrinogen and prothrombin. Neither heparin nor oral anticoagulants interfere with the ECA. Copyright 2004 S. Karger AG, Basel.