FV-TFPI interaction. (A,B) Plasma FV immunodepletion. Pooled plasma from 2 healthy donors was incubated with protein A sepharose beads bearing an anti-FV monoclonal antibody (treated plasma; triangles) or no antibody (control plasma; circles). At regular intervals, subsamples were taken and assayed for FV and full-length TFPI antigen as described in “Plasma FV immunodepletion.” FV levels (A) and TFPI levels (B) were expressed as percentage of the average control plasma level. The experiment was conducted at low (citrated plasma; closed symbols, solid lines) and normal (hirudin/TAP/CTI-anticoagulated plasma; open symbols, dashed lines) levels of free calcium ions. (C,D) Measurement of FV-TFPI interaction by SPR. A CM5 chip with immobilized TFPI was perfused with increasing concentrations of FV as described in “Surface plasmon resonance analysis.” (C) Sensograms recorded at 3400 RU of immobilized TFPI with the indicated FV concentrations. (D) Dose-response curves showing the equilibrium binding response (RU_{240 seconds}) as a function of the FV concentration at 1500, 3400, and 7000 RU of immobilized TFPI.