The GCaMP-R Family of Genetically Encoded Ratiometric Calcium Indicators

Jung Hwa Cho, Carter J. Swanson, Jeannie Chen, Ang Li, Lisa G. Lippert, Shannon E. Boye, Kasey Rose, Sivaraj Sivaramakrishnan, Cheng Ming Chuong, Robert H. Chow

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19 Scopus citations

Abstract

We report on GCaMP-Rs, a new family of genetically encoded ratiometric calcium indicators that extend the virtues of the GCaMP proteins to ratiometric measurements. We have engineered a tandem construct of calcium-dependent GCaMP and calcium-independent mCherry fluorescent proteins. The tandem design assures that the two proteins localize in the same cellular compartment(s) and facilitates pixelwise ratiometric measurements; however, Förster resonance energy transfer (FRET) between the fluorophores reduces brightness of the sensor by up to half (depending on the GCaMP variant). To eliminate FRET, we introduced a rigid α-helix, the ER/K helix, between GCaMP and mCherry. Avoiding FRET significantly increases the brightness (notably, even at low calcium concentrations), the signal-to-noise ratio, and the dynamic range.

Original languageEnglish (US)
Pages (from-to)1066-1074
Number of pages9
JournalACS Chemical Biology
Volume12
Issue number4
DOIs
StatePublished - Apr 21 2017

Bibliographical note

Funding Information:
This research was supported by NIH grant R01 GM85791 to R.H. Chow, R01 EY022931 (to J. Weiland and R.H. Chow), EY12155 EY027193 (to J. Chen), 1DP2 CA186752-01 (to S. Sivaramakrishnan) R01 AR47364, and AR60306 (to C.-M. Chuong).

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