Cytochrome P450 Assay Details
For screening of CYP450, we use the Vivid CYP450 Screening Kits (by ThermoFisher) which enables the rapid measurement of cytochrome P450 enzyme activity using a fluorescent assay. Test compounds are analyzed by their capacity to inhibit the production of a fluorescent signal in reactions using CYP-containing baculosomes and CYP-specific substrates. The cleavage of the fluorogenic substrate results in a fluorescent metabolite. Baculosomes are microsomes prepared from insect cells infected with recombinant baculovirus expressing a human CYP450 isoform.
CYP19A activity is measured with the Aromatase (CYP19A) Inhibitor Screening Kit (BioVision) which is a fluorescence-based assay. The assay uses recombinant human CYP19A and a fluorogenic substrate resulting in a fluorescent metabolite that can be quantified by a standard fluorescence reader.
Setups: Single-dose screening in duplicates or IC50 value determination with 5 or 10 concentrations. Other screening formats are available upon request.
Controls: No inhibitor (DMSO vehicle) control and for every assay, one target-specific control compound is tested in 10-dose IC50 format.
Turnaround time: 10 business days for standard projects. Expedited scheduling and data delivery can be arranged prior to the commencement of the studies.
Report: The raw data, % enzyme activity and control compound IC50 values will be reported in Excel format for single-dose assays. For IC50 orders, raw data, IC50 values, and curve fitting will be delivered in Excel format. Assay conditions, target, and substrate information are available upon request. Requirements for this information should be noted prior to the commencement of the study.
Screening facility: This assay is performed at our screening facility in Malvern, PA, US.
Compound requirements: In brief, for a standard project, 20 µl of a 10 mM DMSO stock or solid material is needed. Less material is needed for large scale screening. Please refer to our FAQs for information regarding compound preparation and shipping.