CYP2B6 and CYP3A4 inhibition

CYP2B6- and CYP3A4-transfected baculosomes from Vivid® CYP450 Blue and Red screening kits, respectively, were obtained from Life Technologies (Carlsbad, CA, USA). Potential inhibitors (and substrates) were screened as described in the Vivid® CYP450 Screening Kits User Guide.

Decreased fluorescence due to chemical inhibition was measured on a Gen5 microplate reader (Synergy H1 Hybrid Reader, BioTek, Winooski, Vermont, USA) at 415/460 nm excitation/emission (CYP2B6) and 550/590 excitation/emission (CYP3A4) at automatic read intervals of 50 seconds for a 30-minute duration in kinetic assay mode. Results were exported into Microsoft Excel, where initial fluorescence was subtracted from fluorescence within the linear part of the curve to calculate change over time and averaged between replicates (n = 3-6).