It is our intent to prepare and use monoclonal antibodies (MAb) for: I) studying structure, size and orientation of epitopes on fibrinogen domains with emphasis on those epitopes whose expression may be altered during/after the fibrinogen-to-fibrin transition; II) identifying neoepitopes on fibrinogen/fibrin fragments and evaluating their clinical significance; III) studying in vivo fibrinogen/fibrin proteolysis in patients where thrombosis is imminent or manifest, already available MAb will be used in these studies; IV) developing immunoassays for measuring fibrinogen/fibrin proteolysis in laboratory animals that are currently used as models for studying thrombogenicity of blood products or efficacy of certain reagents which are thought to be useful for controlling thrombotic disorders. Antisera to fibrinogen or its fragments have provided significant information towards our understanding of the structure-function relationship of this protein and have also been used as clinical reagents. Until recently, such antibodies have been produced by conventional immunization and, as a result, some problems have been encountered. The development of the hybridoma technique by Kohler and Milstein should allow refinement in fibrinogen/fibrin immunoassays. We have now prepared a number of MAb to different regions of the NH2-terminal domain of both fibrinogen and fibrin. Some of these antibodies may become useful in clinical studies. For example, the antibody MAb/1-8C6 is to an epitope in or around the thrombin-susceptible BBeta 14 Arg-15 Gly bond and reactivity is completely abolished when this bond is cleaved. This antibody reacts with peptides released from fibrinogen or fibrin I (a des-FPA fibrin) when either is digested with plasmin. Another MAb (MAb/T2Gls) reacts with fibrin derivatives which contain a neoepitope that is generated as consequence of thrombin cleavage of the BBeta 14 Arg-15 Gly bond. Since the latter MAb does not react with intact fibrinogen, it may be used to identify fibrin II (a des FPA and FPB fibrin) or its degradation products in clinical blood samples. Attempts will now be made to prepare MAb to other neoepitopes. Both in vivo and in vitro immunization protocols will be tried using lipopolysaccharide (LPS) conjugates of fibrinogen/fibrin fragments as immunogens. Preliminary experiments have given promising results. The long-term goals of this research is to prepare MAb to specific epitopes on fibrinogen with the hope that such efforts provide probes for studying the fibrinogen-to-fibrin transition as well as useful clinical reagents.