| Assay Method Information | |
| | Inhibitory Activity Assay |
| Description: | Human liver microsome solution prepared according to the following table (diluted with potassium phosphate solution) Protein con. of working Final protein con. P450 subtype solution (mg/ml) (mg/ml) 1A2 0.133 0.10 2C9 0.0668 0.05 2C19 0.266 0.20 2D6 0.133 010 3A4 0.0668 0.05 3A4 0.133 0.10 150 uL 0.316 mg/mL human liver microsome working solution was added to each well of a 96-well deep plate, and 2 uL of 1 mM specific inhibitor or test compound solution was added to the corresponding wells, and then added to each well 50 uL preheated working solution of each substrate (in 4 mM NADPH solution). After incubating for 5-40 minutes respectively (different enzymes have different incubation times), 200 uL of pre-cooled reaction stop solution was added. All samples were centrifuged at 4000 rpm for 20 minutes. 150 uL of the centrifugal supernatant was then added to a new 96-well deep plate, and added to each well 150 uL of 0.1% formic acid aqueous solution, for analysis by LC/MS/MS (SCI AP4000). |
| Affinity data for this assay | |
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