Protein construct Factor Xa purchased from R&D Systems- Source : Sf9 derived, Met1-Lys488, C-terminal 10-His tag Reconstituted purchased 25 ug to 100 ug/mL in 25 mM MES, 150 mM NaCl, 5 mM CalCl2, pH 6.0 http://rndsystems.com/products/ACFP1063Compounds The 55 compounds were obtained from the K4DD collaboration partners: Sheraz Gul European Screening Port GmbHBuffers Coupling buffer : 10 mM sodium acetate, pH 5.0 Immobilisation : HBS- N, pH 7.5 10 mM HEPES, 150 mM NaCl Assay running buffer : HBS-NP + 2 mM CaCl2 + 2% DMSO 20 mM HEPES, 150 mM NaCl, 2 mM CaCl2, 0.05% Tween 20, 2% DMSO Sample buffer : Identical to assay running buffer Regeneration solution : identical to assay running bufferKinetic titration experiments of 10 ug/ mL FXa immobilised onto a CM5 chip using standard amine coupling chemistry for use with SPR at 25C.Compounds assayed against the surface in a 1:3 dilution series in 2% DMSO