DDE COMPLEX DETERMINATION AS A SPECIFIC MARKER FOR FIBRINOLYSIS

Monoclonal antibodies (McAb) reacting with fibrin degradation products (FbDP), but not with fibrinogen have been produced in order to determine specifically FbDP directly in plasma. Most of the McAb available however do also react with fragment D. Our anti D neo McAb reacts about 10 times less with fragment D than with FbDP but does not react with fibrinogen, fragment X or Y.In clinical investigation, even in pathological conditions in which there is a great release of tissue-type plasminogen activator (tpA), we have shown that fragment D is not generated in patients plasma. Therefore, the reactivity of our McAb with fragment D did not alter the specificity of FbDP assay.On the contrary, using polyacrylamide gel electrophoresis in the presence of SDS followed by immunorevelation, we have evidenced that fragment D is generated in patients undergoing thrombolytic therapy even with rtpA. Therefore, using conventional Elisa procedure (capture of FbDP on polystyrene-immobilized anti D neo antibody and detection by peroxidase-labelled anti fragment D immunoglobulins), the presence of fragment D in patients plasma leads to an overevaluation of FbDP. To avoid this overestimation we have modified the Elisa procedure. The structure of FbDP was taken into acount in order to render the technique specific of FbDP. In FbDP fragment D coming from one fibrin monomer is always associated with fragment E from another fibrin monomer, as DDE complex for example. Therefore after capture of fragment D by the polystyrene-bound anti D neo McAb, FbDP were specifically revealed by peroxidase-labelled anti E antibody (polyclonal or monoclonal anti E may be used). For this reason, this test was named DDE determination and DDE determination can be used in any circumstances to evaluate fibrin degradation.