GRAB(ACh2.0) is a genetically encoded fluorescent sensor for acetylcholine that can be used for real-time detection of endogenous acetylcholine dynamics. It is a G-protein-coupled receptor activation-based ('GRAB') sensor derived from the acetylcholine receptor encoded by the Homo sapiens CHRM3 gene (HGNC:1952); the third intracellular loop (ICL3) of the CHRM3 gene has been replaced with the shorter ICL3 sequence from the Homo sapiens ADRB2 gene (HGNC:286) (to avoid any potential problems with expression and trafficking of the protein) and a circularly permuted form of EGFP (cpEGFP) has been inserted into this ICL3 loop. GRAB(ACh2.0) was derived from GRAB(ACh1.0), by introducing a number of mutations that enhance the dynamic range. Acetylcholine-dependent conformational changes in the receptor result in a change in fluorescence of the conformationally sensitive cpEGFP; GRAB(ACh2.0) shows an increase in fluorescence in response to extracellular acetylcholine (FBrf0240517).

GRAB(ACh2.0) shows weak coupling to downstream signaling pathways in cultured cells, but this coupling had no detectable effect on basic membrane or synaptic responses or on cholinergic transmission in rodent neurons in vitro and in vivo, or on cholinergic responses in Drosophila in vivo (from FBrf0240517).