The development of assays for the detection of fibrin(ogen)olysis based on COOH-terminal A alpha chain epitopes

The COOH-terminal two-thirds of the fibrinogen A alpha chain is a substrate for both factor XIIIa and plasmin and is, therefore, a source of structural markers for the clinical detection of fibrin(ogen)olysis. Monoclonal antibodies (MoAbs) that bind to epitopes within this region (F-102, A alpha 563–578; F-103, A alpha 259–276) have been applied towards the development of two sensitive and specific enzyme-linked immunosorbent assays (ELISAs). The first assay, a capture (F-102)-tag (F-103) ELISA, measures plasma fibrinogen molecules whose A alpha chains are intact. The second assay, a solution phase competitive ELISA based on MoAb F-102, quantifies circulating COOH-terminal A alpha chain degradation products (A alpha FDPs), among the earliest peptides released from fibrinogen during plasmin-mediated fragment X formation. This assay features a novel preliminary plasma absorption step on concanavalin A to recover A alpha FDPs (if present in the sample) in a milieu free of immunologically cross-reactive fibrinogen. Both ELISAs use highly purified fibrinogen as the assay standard for quantitation. In control plasmas, circulating A alpha FDPs accounted for less than 2% of their respective intact fibrinogen A alpha chain concentration, suggesting a physiologic low level of proteolysis occurring at the extreme COOH-terminal portion of the molecule. Plasma A alpha FDPs were elevated (2.3% to 7.8% of their respective intact fibrinogen A alpha chain concentration) in a group of plasma from patients with documented, high serum FDPs (21 to 41 micrograms/mL). Application of the two ELISAs to characterize the course of A alpha chain proteolysis during thrombolytic therapy (TIMI phase 1) indicated that A alpha FDPs were a very early marker of the lytic state (detectable 15 minutes after treatment had been initiated), and that streptokinase and recombinant tissue plasminogen activator appeared to produce significantly different A alpha chain degradation profiles.