Objective

Early assessment of cytochrome P450 (CYP) inhibition has become an essential component of drug discovery screening. The CYP3A4 enzyme is the most abundant enzyme in the liver and plays a major role in metabolizing many xenobiotics in humans. The Promega P450-Glo™ CYP3A4 Assay (Luciferin-IPA) offers an extremely sensitive, high throughput and specific luminescence assay for the examination of CYP3A4 inhibition with pooled Human Liver Microsomes. The Echo 555 liquid handler provides precise and accurate acoustic transfer free of cross-contamination risk. Here, we demonstrate optimization of the P450-Glo™ Assay with Human Liver Microsomes (HLM) in low volume format using the Echo 555 liquid handler. We show that the Echo 555 liquid handler can transfer microsomes, inhibitors and substrate in nanoliter volumes, providing a sensitive and robust small scale assay. IC50 results for four known CYP3A4 inhibitors using our miniaturized assay compared favorably to previously reported values in the literature.