| Assay Method Information | |
| | Determination of FXIIa Inhibition |
| Description: | In vitro inhibition of Factor XIIa was determined using an IC50 assay based on standard literature methods (see e.g Baeriswyl et al., ACS Chem. Biol., 2015, 10 (8) 1861; Bouckaert et al., European Journal of Medicinal Chemistry 110 (2016) 181). Human Factor XIIa (Enzyme Research Laboratories) was incubated at 25° C. with the fluorogenic substrate H-DPro-Phe-Arg-AFC (Peptide Protein Science) and various concentrations of the test compound. Protease activity was measured by monitoring the accumulation of liberated fluorescence from the substrate over 5 min at 25° C. The linear rate of fluorescence increase per minute was expressed as percentage (%) activity. The Km for the cleavage of the substrate by FXIIa was determined by standard transformation of the Michaelis-Menten equation. The compound inhibitor assays were performed at substrate Km concentration. IC50 values were calculated as the concentration of inhibitor giving 50% inhibition (IC50) of the uninhibited enzyme activity (100%). |
| Affinity data for this assay | |
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