P450 glo cyp3a4

P450-Glo CYP3A4, CYP1A2, CYP2B6, and CYP2D6 assay kits were purchased from Promega Corporation. Spheroid 96-well microplates used for the growth of 3D cell spheroid cultures and the black wall/clear bottom 96-well plates were purchased from Corning Life Sciences. The positive controls for induction assays were purchased from Sigma-Aldrich. SH-SY5Y and neural stem cells were plated at 30,000 per well in $80\mathrm{\mu }\mathrm{L}$ of the culture medium in black wall/clear bottom 96-well or spheroid 96-well plates. The assay plates were incubated at 37°C for 48 h. For induction assays, $10\mathrm{\mu }\mathrm{L}$ of omeprazole (CYP1A2) or rifampicin (CYP2B6, CYP2D6, and CYP3A4) with eight concentrations ranging from $21\text{\hspace{0.17em}nM}$ to $46\mathrm{\mu }\mathrm{M}$ were transferred to the assay plates. After the assay plates were incubated at 37°C for 24 h, $10\mathrm{\mu }\mathrm{L}$ P450-Glo substrates [ $3\mathrm{\mu }\mathrm{M}$ Luc-IPA (3A4); $10\mathrm{\mu }\mathrm{M}$ Luc-ME EGE (2D6); $6\mathrm{\mu }\mathrm{M}$ Luc-1A2; $3\mathrm{\mu }\mathrm{M}$ Luc-2B6] were added to assay wells. The assay plates were incubated at 37°C for an additional 1 h. The reactions were stopped by the addition of $100\mathrm{\mu }\mathrm{L}$ P450-Glo detection reagents for each assay. After 20 min incubation at room temperature, the luminescence intensity was quantified using ViewLux™ plate reader. Data were expressed as relative luminescence units. Each data point represents the $\text{mean}\pm \text{standard}\text{\hspace{0.17em}deviation\hspace{0.17em}}\left(\text{SD}\right)$ of three experiments.