| Assay Method Information | |
| | Inhibition Assay |
| Description: | This assay was performed in a 200 l volume in 96-well microtiter plates using cDNA-expressed human hepatic CYP3A4 (supersome, BD Gentest #456202). As a substrate for CYP3A4, 7-benzyloxy-4-trifluoromethyl-coumarin (BFC) was used. The compounds of Examples 1 to 5 and the substrate BFC were dissolved in 100% acetonitrile. The final volume of acetonitrile of the incubation mixture was less than 1% (volume/volume). A potassium phosphate buffer (pH 7.4, final concentration 0.1M), MgCl2 (final concentration 8.3 mM), EDTA (final concentration 1.67 mM), an inventive compound stock solution, CYP3A4 supersome, and NADPH (final concentration 0.25 mM) were added to each well. The reaction was initiated by adding the substrate (BFC, final concentration 30 M) at 37. The incubation was performed for 20 minutes and then the reaction was terminated by the addition of 75 ul of acetonitrile: 0.5 M tris-base=4:1 (volume/volume).Then, a fluorescent signal was measured using a fluorometer. |
| Affinity data for this assay | |
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