| Automation of the Luminescent Based P450-Glo CYP3A4 and 2C9 Inhibition Assay with Human Liver Microsomes |
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Presentation: P03001 Session: Robotic Screening & Automation Technologies - Poster SessionGisela Backfisch and Susann Brixner, Abbott GmbH & Co KG, Ron Aoyama and Steve Morfitt, Abbott Laboratories, Brad Larson and Tracy Worzella, Promega Corporation Presenting Author: Gisela Backfisch, Abbott GmbH & Co. KG - Germany A high-throughput method was automated for screening inhibitions of CYP3A4 and 2C9 in human liver microsomes. To enhance throughput and save valuable working time the luminescent based inhibition assays for CYP3A4 and 2C9 were transferred onto a Tecan Freedom EVO 200. Human liver microsomes were used instead of expressed human CYP3A4 and 2C9 which demands a highly CYP-specific metabolism of the substrates.. The range from 9 nM up to 20 µM was done on one serial dilution plate in a 96-well format. Then the test compounds and the controls were transferred to 2 different plates for the determination of the potential inhibition of CYP3A4 and 2C9. After incubation, the P450-Glo substrates were added and the occurring luminescent signal was measured on a luminescent plate reader. Calculation of IC50s was performed using GraphPadPrism. The results from the automated assays were in good agreement with the results from manual incubations. |