Monoclonal antibodies against antithrombin III and their application in an immunoradiometric assay.

Monoclonal antibodies (MAbs) to antithrombin III (AT III) have been produced and characterized, and a two-site immunoradiometric assay (IRMA) developed, using one of the MAbs as a 'catcher' antibody and a radiolabelled affinity-purified rabbit antibody for detection. The IRMA could be performed in one day and the optimum concentration range of AT III was 1-100 ng/ml. Assays on AT III concentrates by the IRMA method showed a good correlation with AT III antigen values measured by the immunoelectrophoresis method (Laurell), the IRMA results averaging 94.3% of the Laurell values. Two of the MAbs (11 and 16) inhibited heparin cofactor but not AT III progressive activity, and the binding of AT III to MAb 11, used for the IRMA, was blocked by heparin. These results indicate that MAb 11 is directed at or near to the heparin binding site, and could therefore be useful in the study of structural aspects of this site in normal and genetically abnormal AT III.