[Functional characterization of a monoclonal antibody which interferes with the binding of t-PA to fibrin].
OBJECTIVE
Development of monoclonal antibodies capable of inhibiting the specific binding of t-PA to fibrin.


METHODS
After immunization of Balb/c mice with recombinant t-PA (rt-PA) we selected the monoclonal antibody MA3B5 by its ability to inhibit the binding of t-PA to fibrin, and the MA2C1 devoid of this property. The influence of such antibodies was evaluated on fibrin plates, amidolytic assays and clot lysis assays. Furthermore, their interference with the activator bound to fibrin was assayed with a spectrophotometric solid phase assay (SOFIA).


RESULTS
The results showed that MA3B5 totally inhibited the t-PA induced fibrinolytic activity on fibrin plates, reduced amidolytic activity by 86.5% and inhibited the clot lysis induced by t-PA in a dynamic system. In contrast, the MA2C1 showed no inhibition. By assessing the binding of t-PA to fibrin with the SOFIA assay we could demonstrate that only the MA3B5 reduced significantly (up to 90% with 100 micrograms/mL of antibody) the amount of t-PA bound to fibrin surface.


CONCLUSIONS
We have purified and characterized a monoclonal antibody which specifically blocks the fibrin binding site of t-PA.
Orbe
J
J
Laboratorio de Pared Vascular y Trombosis, Facultad de Medicina, Universidad de Navarra, Pamplona.
Montes
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R
Chordá
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C
Páramo
J A
JA
Rocha
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E
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D005260:Female; D005337:Fibrin / Q000378:metabolism*; D005342:Fibrinolysis / Q000187:drug effects*; D051379:Mice; D008807:Mice, Inbred BALB C; D009842:Oligopeptides / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011994:Recombinant Proteins / Q000276:immunology; D010959:Tissue Plasminogen Activator / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D009842:Oligopeptides; D011994:Recombinant Proteins; C018788:valyl-leucyl-lysine 4-nitroanilide; D005337:Fibrin; D010959:Tissue Plasminogen Activator
Caracterización funcional de un anticuerpo monoclonal que interfiere la unión del t-PA a la fibrina.