A Real‐Time, Plate‐Based BRET Assay for Detection of cGMP in Primary Cells

Abstract

Cyclic guanosine monophosphate (cGMP) is a second messenger involved in the regulation of numerous physiological processes. The modulation of cGMP is important in many diseases, but reliably assaying cGMP in live cells in a plate‐based format with temporal resolution is challenging. The Förster/fluorescence resonance energy transfer (FRET)‐based biosensor cGES‐DE5 has a high temporal resolution and high selectivity for cGMP over cAMP, so we converted it to use bioluminescence resonance energy transfer (BRET), which is more compatible with plate‐based as-says. This BRET variant, called CYGYEL (cyclic GMP sensor using YFP‐PDE5‐Rluc8), was cloned into a lentiviral vector for use across differen..