Summary FlipGFP assay characterizes the intracellular drug target engagement to M pro and PL pro and can be performed in the biosafety level 1/2 settings. Here, we provide the detailed protocol for the cell-based FlipGFP assay to identify and characterize SARS-CoV-2 M pro and PL pro inhibitors. We describe steps for cell passage and seeding, transfection, addition of compounds, and their incubation and timing. We then detail the quantification of the fluorescence signal of the assay For complete details on the use and execution of this protocol, please refer to Ma et al. 1 Graphical abstract Highlights • Cell based assay for the characterization of SARS-CoV-2 M pro and PL pro inhibitors • Predict antiviral activity of protease inhibitors at biosafety level 1/2 laboratory • Reveal intracellular drug target engagement, cytotoxicity, and membrane permeability Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. FlipGFP assay characterizes the intracellular drug target engagement to M pro and PL pro and can be performed in the biosafety level 1/2 settings. Here, we provide the detailed protocol for the cell-based FlipGFP assay to identify and characterize SARS-CoV-2 M pro and PL pro inhibitors. We describe steps for cell passage and seeding, transfection, addition of compounds, and their incubation and timing. We then detail the quantification of the fluorescence signal of the assay.
【저자키워드】 Microbiology, Chemistry,