A catching ELISA has been developed that permits a quantitation of the anticoagulant hirudin in buffer, urine and plasma. In plasma hirudin can be determined in concentrations ranging from 0.2 to 25 ng/ml (2.4 X 10(-3) to 0.3 AT-U/ml), in urine between 0.8 and 200 ng/ml (0.01 and 2.4 AT-U/ml). The enzyme immunoassay allows a rapid, sensitive and reproducible quantitation of hirudin, and can thus be used to assess the pharmacokinetics of the anticoagulant in patients after parenteral and/or topic administration.