MONOCLONAL-ANTIBODY DIRECTED TO A FIBRINOGEN A-ALPHA-NUMBER-529-539 EPITOPE INHIBITS ALPHA-CHAIN CROSS-LINKING BY TRANSGLUTAMINASES

A monoclonal antibody (5A2) recognizing a segment near the C-terminus of the fibrin(ogen) A alpha-chain (Act #529-539) was found to inhibit a-chain crosslinking catalyzed by coagulation factor XIII(a) and by ti ssue-transglutaminase. The rapid gamma-chain cross-linking by factor X III(a) was not affected by the antibody. Results obtained from direct binding and competitive immunoassay established that the antigenic det erminant recognized by 5A2 was included within the CNBr fragment refer red to as CNBr X (A alpha #518-584), and that it survived trypsin dige stion but was destroyed by treatment with Staph V-8 protease or chymot rypsin. Reverse-phase (C-18) high performance liquid chromatography (H PLC) was employed to obtain a CNBr X tryptic fingerprint, which was su bsequently characterized by compositional and NH2-terminal analysis. A ssay of the HPLC column effluent revealed a single peak of 5A2 immunor eactivity that coincided with elution of the eleven-residue tryptic pe ptide, A alpha #529-539. When this isolated peptide and its parent CNB r fragment were employed as solution phase competitors in the 5A2 immu noassay, the relative cross-reactivities (18.3%, peptide: fragment) in dicated that a significant proportion of the 5A2 epitope was preserved within the small peptide. This is a region that is released from fibr inogen early in its degradation by plasmin. Thus, the antibody can be used as a probe for intact fibrin(ogen) and C-terminal (A)alpha-chain fragments, in addition to assessing roles of the A alpha-chain C-termi nus in cross-linking.