Ligand blotting of PS and FXa to TFPI. TFPI proteins were electrophoresed on 12% SDS-PAGE gel and transferred to a nitrocellulose membrane. The amount of each protein that was electrophoresed and blotted is labeled at the top. (A-B) Binding of PS to TFPI. PS was used to probe the membrane, and the bound PS was detected by an anti-PS polyclonal antibody as described in “Ligand and Western blotting.” (C-D) Binding of FXa to TFPI. After PS detection in panels A and B, the membranes were stripped with 0.1mM glycine, pH 2.2, and reprobed with FXa. The bound FXa was then detected using a goat anti-FX polyclonal antibody as described in “Ligand and Western blotting.” The positions of molecular weight markers are indicated on the left.