Absolute measurement of cellular activities using photochromic single-fluorophore biosensors and intermittent quantification.

ABSTRACT: Genetically-encoded biosensors based on a single fluorescent protein are widely used to visualize analyte levels or enzymatic activities in cells, though usually to monitor relative changes rather than absolute values. We report photochromism-enabled absolute quantification (PEAQ) biosensing, a method that leverages the photochromic properties of biosensors to provide an absolute measure of the analyte concentration or activity. We develop proof-of-concept photochromic variants of the popular GCaMP family of Ca²⁺ biosensors, and show that these can be used to resolve dynamic changes in the absolute Ca²⁺ concentration in live cells. We also develop intermittent quantification, a technique that combines absolute aquisitions with fast fluorescence acquisitions to deliver

SUBMITTER: Bierbuesse F 

PROVIDER: S-EPMC8986857 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature