Novel cell-based assays employing voltage and calcium dyes
AssigneeAtto Bioscience, Inc.
InventorsYong Yao, Liang Cao, Jianming Lu, Isabel Llorente
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Abstract
Disclosed are novel compositions and methods employing cyclic nucleotide-gated channels and fluorescence dyes and other indicators in detecting changes of intracellular cAMP levels, for instance in response to the stimulation of G-protein-coupled receptors. CNG mutants comprising one or more pore mutations that enhance sensitivity to cAMP and decrease divalent cation-mediated blockage are used in a variety of assays, including detection of the G-protein-coupled receptor activity. Also disclosed are novel dye quencher formulations that are particularly suited for cell-based assays of protein interactions.
Claims24
An isolated polynucleotide comprising a sequence encoding a mutant cyclic nucleotide-gated (CNG) channel comprising:
- (a) at least one mutation that decreases divalent cation-mediated blockage of cation flux;
- and
- (b) at least one mutation that makes the channel more sensitive to cAMP than a channel that does not comprise the mutation.
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Dependencies are inferred from the claim text.