Binding of fluorescein isothiocyanate-labeled alpha-thrombin to peritoneal mast cells
Strukova, S.M.; Khlgatian, S.V.
Thrombosis Research 64(6): 795-796
1991
ISSN/ISBN: 0049-3848 PMID: 1798971 Document Number: 379306
The interaction of bovine .alpha.-thrombin with peritoneal mast cells was studied using FITC-labeled enzyme. Thrombin was modified with FITC in the presence of heparin and was separated from heparin and free FITC by gel-filtration at HPLC yielding FITC-labeled .alpha.-thrombin with intact additional recognition binding site for high molecular substrates and cell receptors. Equilibrium studies have shown that the binding of thrombin to peritoneal mast cells is active independent, rapid, specific, saturable and reversible. Equilibrium between bound and free thrombin is attained within I min and Scatchard analysis indicates a population of approximately 54 .times. 103 sites/cell with a dissociation constant of 1.3 .times. 10-9 M. FITC-labeled .alpha.-thrombin binds to peritoneal mast cells in a temperature-dependent manner with optimum at 37.degree. C. These results indicate that FITC-labeled .alpha.-thrombin binds to peritoneal mast cells with high affinity.