Application of bioluminescence resonance energy transfer to quantitate cell-surface expression of membrane proteins.

We report a bioluminescence resonance energy transfer (BRET) assay to quantitate the fraction of an engineered membrane protein at the cell surface versus inside the cell. As test cases, we engineered two different G protein-coupled receptors (GPCRs) in which a NanoLuc luciferase (NLuc) and a HaloTag are fused to the extracellular amino-terminal tail of the receptors. We then employed a pulse-chase labeling approach relying on two different fluorescent dyes with distinctive cell permeability properties.