Assay Method Information | |
| In Vitro Inhibition Assay |
Description: | The inhibitory activity of the compounds of the invention was demonstrated in vitro by measuring the inhibition of recombinant human Cathepsin S as follows: To a 384 well microtitre plate is added 10 μl of a 100 μM solution of test compound in assay buffer (100 mM sodium acetate pH5.5, 5 mM EDTA, 5 mM dithiothreitol) with 10% dimethylsulfoxide (DMSO), plus 20 μl of 250 μM solution of the substrate Z-Val-Val-Arg-AMC (7-amido-coumarine derivative of the tripeptide N-benzyloxy-carbonyl-Val-Val-Arg-OH) in assay buffer and 45 μl of assay buffer. 25 μl of a 2 mg/l solution of activated recombinant human cathepsin S, in assay buffer, is then added to the well, yielding a final inhibitor concentration of 10 μM.Enzyme activity is determined by measuring the fluorescence of the liberated aminomethylcoumarin at 440 nM using 390 nM excitation, at 20 minutes. |
Affinity data for this assay | |
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