A near-infrared genetically encoded calcium indicator for in vivo imaging

Anton A. Shemetov, Mikhail V. Monakhov, Qinrong Zhang, Jose Ernesto Canton-Josh, Manish Kumar, Maomao Chen, Mikhail E. Matlashov, Xuan Li, Wei Yang, Liming Nie, Daria M. Shcherbakova, Yevgenia Kozorovitskiy, Junjie Yao, Na Ji, Vladislav V. Verkhusha

Research output: Contribution to journalArticlepeer-review

79 Scopus citations


While calcium imaging has become a mainstay of modern neuroscience, the spectral properties of current fluorescent calcium indicators limit deep-tissue imaging as well as simultaneous use with other probes. Using two monomeric near-infrared (NIR) fluorescent proteins (FPs), we engineered an NIR Förster resonance energy transfer (FRET)-based genetically encoded calcium indicator (iGECI). iGECI exhibits high levels of brightness and photostability and an increase up to 600% in the fluorescence response to calcium. In dissociated neurons, iGECI detects spontaneous neuronal activity and electrically and optogenetically induced firing. We validated the performance of iGECI up to a depth of almost 400 µm in acute brain slices using one-photon light-sheet imaging. Applying hybrid photoacoustic and fluorescence microscopy, we simultaneously monitored neuronal and hemodynamic activities in the mouse brain through an intact skull, with resolutions of ~3 μm (lateral) and ~25–50 μm (axial). Using two-photon imaging, we detected evoked and spontaneous neuronal activity in the mouse visual cortex, with fluorescence changes of up to 25%. iGECI allows biosensors and optogenetic actuators to be multiplexed without spectral crosstalk.

Original languageEnglish (US)
Pages (from-to)368-377
Number of pages10
JournalNature biotechnology
Issue number3
StatePublished - Mar 2021

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering


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