Prothrombin activation in rabbits

The suitability of rabbit prothrombin activation fragment F 1.2 as a marker for the activation of the coagulation system was tested. Monoclonal antibo dies to rabbit F 1.2 were raised, and a competitive F 1.2 ELISA was develop ed. Within the detection limit of the ELISA, no increase in rabbit F 1.2 wa s detected upon recalcification of plasma, whereas human F 1.2 increased 15 00-fold. The apparent lack of F 1.2 formation in rabbit serum was confirmed by immunoblotting analysis of endogenous and biotin-labeled prothrombin. M eizothrombin and the B-chain of thrombin were the only prothrombin fragment s detectable. In contrast, labeled human prothrombin formed, in addition, p rethrombin 2 and F 1.2 in both human and rabbit serum. In contrast, rabbit F 1.2 formation could be demonstrated using purified rabbit prothrombin and factor Xa. These observations raise the possibility that rabbit prothrombi n is less susceptible than the human counterpart to factor Xa cleavage at t he 271/272 peptide bond. Thus, the primary structure of rabbit prothrombin was deduced by cDNA sequencing. While the 320/321 Xa cleavage site giving r ise to meizothrombin was identical in rabbit and human prothrombin, the fla nking region of the 271/272 Xa sensitive site contained a six amino acid de letion in the rabbit sequence. Taken together, these observations suggest t hat the observed differences between human and rabbit prothrombin activatio n may be due to different susceptibilities of the two Xa cleavage sites rat her than plasma or serum cofactor(s), (C) 1999 Elsevier Science Ltd.