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Ratiometric Matryoshka biosensors from a nested cassette of green- and orange-emitting fluorescent proteins. | LitMetric

AI Article Synopsis

Article Abstract

Sensitivity, dynamic and detection range as well as exclusion of expression and instrumental artifacts are critical for the quantitation of data obtained with fluorescent protein (FP)-based biosensors in vivo. Current biosensors designs are, in general, unable to simultaneously meet all these criteria. Here, we describe a generalizable platform to create dual-FP biosensors with large dynamic ranges by employing a single FP-cassette, named GO-(Green-Orange) Matryoshka. The cassette nests a stable reference FP (large Stokes shift LSSmOrange) within a reporter FP (circularly permuted green FP). GO- Matryoshka yields green and orange fluorescence upon blue excitation. As proof of concept, we converted existing, single-emission biosensors into a series of ratiometric calcium sensors (MatryoshCaMP6s) and ammonium transport activity sensors (AmTryoshka1;3). We additionally identified the internal acid-base equilibrium as a key determinant of the GCaMP dynamic range. Matryoshka technology promises flexibility in the design of a wide spectrum of ratiometric biosensors and expanded in vivo applications.Single fluorescent protein biosensors are susceptible to expression and instrumental artifacts. Here Ast et al. describe a dual fluorescent protein design whereby a reference fluorescent protein is nested within a reporter fluorescent protein to control for such artifacts while preserving sensitivity and dynamic range.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5585204PMC
http://dx.doi.org/10.1038/s41467-017-00400-2DOI Listing

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