Fibronectin and vitronectin, but not α-2-macroglobulin, inhibit PF4 binding to platelets. GFPs were incubated with 10 µg/mL PF4 in the presence of increasing concentrations of α-2-macroglobulin (A), vitronectin (B), or fibronectin or the respective volumes of its solvent buffer (C). (D) GFPs were incubated with 10 µg/mL PF4 in the presence of 50 µg/mL intact fibronectin (FN), fibronectin gelatin-binding fragment (FN GBF), fibronectin heparin-binding fragment (FN HBF), fibronectin fragments 7-10 of type III domain (FN III 7-10), or fibronectin fragments 12-14 of type III domain (FN III 12-14). (A-D) PF4 binding was detected with an FITC-labeled anti-human PF4 antibody using flow cytometry, and antibody binding was quantified by GMFI. Surface-bound PF4 in the presence of buffer was defined as 100%. Data are mean ± SD of 3 independent experiments. (E) Schematic structure of plasma fibronectin. Fibronectin exists as a dimer, with each monomer composed of 12 type I repeats (circles), 2 type II repeats (hexagons), and 15 type III repeats (diamonds). In contrast to the cellular form, plasma fibronectin lacks the alternatively spliced domains EIIIA and EIIIB (not shown) but includes a variable region (square) in 1 of the subunits. The subunits are linked by 2 disulfide bridges located close to the C termini. Soluble plasma fibronectin occurs as a compact curled-up form and becomes elongated when binding to integrins on cell surfaces. Additional binding sites are indicated for heparin, collagen, gelatin, and fibrin. Brackets denote the fragments used in this study. *P < .05, **P < .001 vs buffer.