title: Coagulation factor IXa binding to activated platelets and platelet-derived microparticles: a flow cytometric study.
abstract: Factor IX plays a central role in blood coagulation, since it can be activated by either XIa (intrinsic pathway) or tissue factor-VIIa (extrinsic pathway). Activated factor IX (IXa), in a surface-bound complex with factor VIIIa, then activates factor X. Platelets provide the catalytic surface upon which this Xase complex is assembled in vivo. We have used flow cytometry to examine binding of factor IXa to thrombin-activated platelets in the absence of added VIIIa. Platelet-bound IXa and platelet protein GPIb were detected by indirect immunofluorescence staining followed by two-color flow cytometric analysis. Microparticles were identified by their light scattering characteristics. Two binding sites for factor IXa were detected. The high affinity binding site saturated at about 10 nM, with a Kd of 1.6 nM. A second binding curve, with a Kd of about 100 nM, was observed at higher concentrations of IXa. The high affinity factor IXa binding sites comprise about 7% of the total factor IXa binding. Binding to both sites was dependent on the presence of calcium. Thus, we conclude that factor IXa, in addition to its high affinity binding, has a calcium-dependent low affinity association with activated platelets and microparticles. Sims et al, have shown that binding sites for a different coagulation factor, factor Va, are concentrated on microparticles relative to platelet membrane proteins, such as GPIb. GPIb is distributed on platelets and microparticle in proportion to plasma membrane surface.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Pathology, University of North Carolina, Chapel Hill.
authors.forename: M
authors.initials: M
authors.lastname: Hoffman
authors.forename: D M
authors.initials: DM
authors.lastname: Monroe
authors.forename: H R
authors.initials: HR
authors.lastname: Roberts
mesh_terms: D001792:Blood Platelets / Q000378:metabolism*; D015949:Factor IXa / Q000378:metabolism*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D011485:Protein Binding; D011869:Radioligand Assay
chemical_list: D015949:Factor IXa