Multiplexed 3D FRET imaging in deep tissue of live embryos

Ming Zhao; Xiaoyang Wan; Yu Li; Weibin Zhou; Leilei Peng

doi:10.1038/srep13991

Multiplexed 3D FRET imaging in deep tissue of live embryos

Ming Zhao, Xiaoyang Wan, Yu Li, Weibin Zhou, Leilei Peng

Research output: Contribution to journal › Article › peer-review

22 Scopus citations

Abstract

Current deep tissue microscopy techniques are mostly restricted to intensity mapping of fluorophores, which significantly limit their applications in investigating biochemical processes in vivo. We present a deep tissue multiplexed functional imaging method that probes multiple Förster resonant energy transfer (FRET) sensors in live embryos with high spatial resolution. The method simultaneously images fluorescence lifetimes in 3D with multiple excitation lasers. Through quantitative analysis of triple-channel intensity and lifetime images, we demonstrated that Ca 2+ and cAMP levels of live embryos expressing dual FRET sensors can be monitored simultaneously at microscopic resolution. The method is compatible with a broad range of FRET sensors currently available for probing various cellular biochemical functions. It opens the door to imaging complex cellular circuitries in whole live organisms.

Original language	English (US)
Article number	13991
Journal	Scientific reports
Volume	5
DOIs	https://doi.org/10.1038/srep13991
State	Published - Sep 21 2015

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title = "Multiplexed 3D FRET imaging in deep tissue of live embryos",

abstract = "Current deep tissue microscopy techniques are mostly restricted to intensity mapping of fluorophores, which significantly limit their applications in investigating biochemical processes in vivo. We present a deep tissue multiplexed functional imaging method that probes multiple F{\"o}rster resonant energy transfer (FRET) sensors in live embryos with high spatial resolution. The method simultaneously images fluorescence lifetimes in 3D with multiple excitation lasers. Through quantitative analysis of triple-channel intensity and lifetime images, we demonstrated that Ca 2+ and cAMP levels of live embryos expressing dual FRET sensors can be monitored simultaneously at microscopic resolution. The method is compatible with a broad range of FRET sensors currently available for probing various cellular biochemical functions. It opens the door to imaging complex cellular circuitries in whole live organisms.",

author = "Ming Zhao and Xiaoyang Wan and Yu Li and Weibin Zhou and Leilei Peng",

note = "Funding Information: This research was supported by NIH grants to L.P. (R00EB008737 and R01EB015481). W.Z. was supported by grants from NIH (R00DK091405), American Society of Nephrology (Carl W. Gottschalk Research Scholar Grant) and National Kidney Foundation.",

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doi = "10.1038/srep13991",

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AU - Zhao, Ming

AU - Wan, Xiaoyang

AU - Li, Yu

AU - Zhou, Weibin

AU - Peng, Leilei

N1 - Funding Information: This research was supported by NIH grants to L.P. (R00EB008737 and R01EB015481). W.Z. was supported by grants from NIH (R00DK091405), American Society of Nephrology (Carl W. Gottschalk Research Scholar Grant) and National Kidney Foundation.

PY - 2015/9/21

Y1 - 2015/9/21

N2 - Current deep tissue microscopy techniques are mostly restricted to intensity mapping of fluorophores, which significantly limit their applications in investigating biochemical processes in vivo. We present a deep tissue multiplexed functional imaging method that probes multiple Förster resonant energy transfer (FRET) sensors in live embryos with high spatial resolution. The method simultaneously images fluorescence lifetimes in 3D with multiple excitation lasers. Through quantitative analysis of triple-channel intensity and lifetime images, we demonstrated that Ca 2+ and cAMP levels of live embryos expressing dual FRET sensors can be monitored simultaneously at microscopic resolution. The method is compatible with a broad range of FRET sensors currently available for probing various cellular biochemical functions. It opens the door to imaging complex cellular circuitries in whole live organisms.

AB - Current deep tissue microscopy techniques are mostly restricted to intensity mapping of fluorophores, which significantly limit their applications in investigating biochemical processes in vivo. We present a deep tissue multiplexed functional imaging method that probes multiple Förster resonant energy transfer (FRET) sensors in live embryos with high spatial resolution. The method simultaneously images fluorescence lifetimes in 3D with multiple excitation lasers. Through quantitative analysis of triple-channel intensity and lifetime images, we demonstrated that Ca 2+ and cAMP levels of live embryos expressing dual FRET sensors can be monitored simultaneously at microscopic resolution. The method is compatible with a broad range of FRET sensors currently available for probing various cellular biochemical functions. It opens the door to imaging complex cellular circuitries in whole live organisms.

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Multiplexed 3D FRET imaging in deep tissue of live embryos

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