FIGURE 1.
Identification of vWbp ligands isolated from human plasma and comparative analysis with Coa. A, human plasma (500 μl) was flowed over Strep-Tactin resin uncharged (control) or charged with H6vWbpstrep (100 nmol). Bound proteins were eluted by boiling the resin in sample buffer, separated by SDS-PAGE, and visualized after Coomassie staining. The identity of proteins labeled 1–9 was performed by microcapillary LC/MS/MS techniques. A complete list of protein hits can be found in Table 2. PT, prothrombin; FG, fibrinogen. B and C, for the comparative analysis of Coa and vWbp ligands, human plasma (500 μl) was flowed over Strep-Tactin resin uncharged (control) or charged with either H6vWbpstrep (100 nmol) or Coastrep (100 nmol). Bound proteins were eluted by boiling the resin in sample buffer and separated by SDS-PAGE. Proteins in gels were visualized by Coomassie staining (B) or after transfer to PVDF membranes for immunoblotting using specific antibodies against prothrombin, FXIII subunit A (FXIII A), and fibronectin (C).