Antipeptide Monoclonal Antibodies to Defined Fibrinogen A
Chain
Regions: Anti-A 487-498, a Structural Probe for Fibrinogenolysis
Joan H. Sobel,
Ilya Trakht,
Nicolas Pileggi, and
Hong Qi Wu
From the Department of Medicine, College of Physicians & Surgeons of
Columbia University, New York, NY.
The fibrinogen 
C domain (A 220-610) is one of the earliest
targets attacked by plasmin following fibrinolytic system activation. Monoclonal antibodies (MoAbs) to defined sequences within the C
domain provide the opportunity to explore the structure-function relationships involved in plasmin's interaction with its A chain substrate at greater resolution and can serve as reagents with potential clinical use for detecting fibrinogenolysis in vivo. The MoAb
F-104 was raised against a multiple antigenic peptide derivative
modelled after the hydrophilic 12-residue sequence corresponding to
A 487-498 within the C domain. A sensitive solution phase
competitive enzyme-linked immunosorbent assay (ELISA) was developed for
MoAb F-104 that can be applied for the direct measurement of intact
fibrinogen (purified or plasma; ED50%
5 pmol A chain
equivalents/mL), with negligible cross-reactive interference from
peptide cleavage products released by plasmin from the COOH-terminal
end of the A chain (<3%). Immunoblotting and ELISA studies to
characterize the fate of the F-104 epitope during fibrinogenolysis in
vitro indicated a rapid loss of fibrinogen-associated immunoreactivity
that reflected the heterogeneity of plasmin cleavage sites within the
C domain; cleavage at the 493-494 arg-his bond destroyed the F-104
epitope, while cleavage at other sites released it in an altered,
inaccessible, conformation within the structure of 35- to
40-kD and 17.5- to 18-kD A chain degradation products. Application of the F-104 ELISA to monitor the course of A chain proteolysis in a small study population of patients undergoing thrombolytic therapy for myocardial infarction (n = 14) showed that
the loss of fibrinogen-associated F-104 immunoreactivity was a very
early marker (within 15 to 30 minutes) of in vivo fibrinogenolysis. Additional data obtained suggest that MoAb F-104 may have promise as a
reagent for evaluating the creation of an effective lytic state early
during therapy, information that could help determine the need for
further clinical intervention. Thus, these studies illustrate a
rational, targeted, approach towards the development of a novel
antifibrinogen MoAb whose application as a structural probe for the
region A 487-498 in vitro and in vivo can provide new insights into
the various molecular forms of fibrinogen that circulate under
physiologic conditions and in disease.
Blood, Vol. 91 No. 5 (March 1), 1998:
pp. 1590-1598
© 1998 by The American Society of Hematology.
