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ISSN 2410-7751 (Print)
ISSN 2410-776X (Online)

 4 2013

"Biotechnologia Acta" v. 6, no. 4, 2013
https://doi.org/10.15407/biotech6.04.033
Р. 33-42, Bibliography 24, English
Universal Decimal classification: 547.962.4

USAGE OF MONOCLONAL ANTIBODIES FOR DETERMINATION OF LOCALIZATION OF ANTIGENIC DETERMINANTS AND FIBRIN POLYMERIZATION SITES WITHIN FIBRINOGEN AND FIBRIN MOLECULES AND THEIR APPLICATION IN TEST--SYSTEMS FOR DIAGNOSTICS AND THE THREAT OF THROMBUS FORMATION

E. V. Lugovskoi, I. N. Kolesnikova, S. V. Komisarenko

Palladian Biochemistry Institute of National Academy of Sciences of Ukraine, Kyiv, Ukraine

It was shown by monoclonal antibodies that BβN-region of fibrin desA molecule (Bβ1-53) comprises the polymerization site including the peptide bond Bβ14-15. This site participates in the second stage of fibrin polymerization — lateral association of protofibrils. In the Bβ15-53 fragment was also found the site called "C", which together with the site "A" participate in the first stage of polymerization — the protofibrils formation. The model of the primary intermolecular interaction of fibrin was designed. It was found by monoclonal antibodies II-4d the site ("c") in the N-terminal half of γ chain of the fibrin D-region. This site participates in the protofibrils formation and is complement to site "C" as we assume.

We have discovered two neoantigenic determinants. One of these determinants exposes within the coiledcoil fragment Bβ126-135 of fibrin as a result of fibrinopeptide A splitting off from fibrinogen by thrombin. The structural rearrangements discovered in this site of the fibrin molecule are necessary for the following protofibrils lateral association. The second neoantigenic determinant is localized in the fragment Bβ134-190 of D-dimer formed after plasmin degradation of fibrin stabilized by FXIIIa. We have obtained the fibrin-specific monoclonal antibodie FnI-3C to the first determinant and D-dimer-specific mAb III-3b to the second one.

Three monoclonal antibodies were obtained against the αC-region of fibrin(ogen) molecule. It has been experimentally shown by of one of them that αC-domains is connected with the fibrinopeptides B in fibrinogen and fibrin desA molecules, but removes from the core of the molecules after fibrinopeptides B splitting off by thrombin. Two other monoclonal antibodies specifically inhibit the fibrin polymerization by blocking two unknown polymerization sites within the αC-region.

The test-systems for the soluble fibrin and D-dimer quantification in human blood plasma were designed on the basis of monoclonal antibodies FnI-3C and III-3b as "catch"-antibodies and one II-4d as a "tag"-antibody, respectively. The clinical trials of the test-systems were carried out in Ukraine. It was shown that for the prediction of postoperative thrombotic complications and monitoring the efficiency of antithrombotic therapy the simultaneous quantification of soluble fibrin and D-dimer before the operation and at different time intervals after the operation is required. Only in this case it is possible to get information about the state of the balance between blood coagulation and fibrinolytic systems, and determine the degree of the threat of thrombosis.

Key words: fibrinogen, fibrin, monoclonal antibodies, thrombosis, diagnostic test systems.

© Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, 2013