Download PDF
Semin Thromb Hemost 1995; 21(2): 184-192
DOI: 10.1055/s-2007-1000394
Copyright © 1995 by Thieme Medical Publishers, Inc.

Development and Validation of Two Enzyme-Linked Immunosorbent Assay (ELISA) Methods for Recombinant Hirudin

Lalitha Iyer* , Mariette Adam† , Jean Amiral, Jawed Fareed* ‡ , Edward Bermes‡  JR. 
  • *From the Department of Pharmacology, Loyola University Medical Center, Maywood, IL,
  • †Serbio Research Laboratory, Gennevilliers, France, and
  • ‡Department of Pathology, Loyola University Medical Center, Maywood, IL.
Further Information

Publication History

Publication Date:
06 February 2008 (online)

  PDF Download  Permissions and Reprints

Abstract

Recombinant hirudin is currently being developed as a potential prophylactic and therapeutic antithrombotic drug in various clinical indications such as angina and deep venous thrombosis. In this report, we have discussed the production of specific polyclonal antibodies to recombinant hirudin (rH) and the development of two ELISA methods to measure rH concentrations in biological fluids: a sandwich and a competitive ELISA method. Intra- and inter-assay variations in the two methods are extremely low (3-7%). The competitive ELISA method is rapid, simple and highly reproducible. Saturation binding curves, selection of appropriate incubation times, recovery of different hirudin variants and reactivity in the presence of thrombin are discussed. The methods can be easily adapted to monitor hirudin concentrations in the clinical laboratory for diagnostic purposes as well as for performing pharmacokinetic studies.

Key words:

Hirudin - recombinant hirudin - ELISA - hirudin assays

      >