TY - JOUR
T1 - The GCaMP-R Family of Genetically Encoded Ratiometric Calcium Indicators
AU - Cho, Jung Hwa
AU - Swanson, Carter J.
AU - Chen, Jeannie
AU - Li, Ang
AU - Lippert, Lisa G.
AU - Boye, Shannon E.
AU - Rose, Kasey
AU - Sivaramakrishnan, Sivaraj
AU - Chuong, Cheng Ming
AU - Chow, Robert H.
N1 - 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).
Publisher Copyright:
© 2017 American Chemical Society.
PY - 2017/4/21
Y1 - 2017/4/21
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=85018480689&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85018480689&partnerID=8YFLogxK
U2 - 10.1021/acschembio.6b00883
DO - 10.1021/acschembio.6b00883
M3 - Article
C2 - 28195691
AN - SCOPUS:85018480689
SN - 1554-8929
VL - 12
SP - 1066
EP - 1074
JO - ACS Chemical Biology
JF - ACS Chemical Biology
IS - 4
ER -