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Biophys J. 2017 Nov 21;113(10):2299-2309. doi: 10.1016/j.bpj.2017.09.007. Epub 2017 Oct 7.

Near-Infrared Fluorescent Proteins Engineered from Bacterial Phytochromes in Neuroimaging.

Author information

1
Media Lab, MIT, Cambridge, Massachusetts; MIT McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts.
2
Media Lab, MIT, Cambridge, Massachusetts; Harvard-MIT Division of Health Sciences and Technology, MIT, Cambridge, Massachusetts.
3
MIT McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts.
4
MIT McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts; Department of Brain and Cognitive Sciences, MIT, Cambridge, Massachusetts.
5
Department of Cell Biology and Neuroscience, Montana State University, Bozeman, Montana.
6
Media Lab, MIT, Cambridge, Massachusetts; MIT McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts; Department of Brain and Cognitive Sciences, MIT, Cambridge, Massachusetts; Department of Biological Engineering, MIT, Cambridge, Massachusetts; MIT Center for Neurobiological Engineering, MIT, Cambridge, Massachusetts. Electronic address: esb@media.mit.edu.
7
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York; Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, Bronx, New York. Electronic address: vladislav.verkhusha@einstein.yu.edu.

Abstract

Several series of near-infrared (NIR) fluorescent proteins (FPs) were recently engineered from bacterial phytochromes but were not systematically compared in neurons. To fluoresce, NIR FPs utilize an enzymatic derivative of heme, the linear tetrapyrrole biliverdin, as a chromophore whose level in neurons is poorly studied. Here, we evaluated NIR FPs of the iRFP protein family, which were reported to be the brightest in non-neuronal mammalian cells, in primary neuronal culture, in brain slices of mouse and monkey, and in mouse brain in vivo. We applied several fluorescence imaging modes, such as wide-field and confocal one-photon and two-photon microscopy, to compare photochemical and biophysical properties of various iRFPs. The iRFP682 and iRFP670 proteins exhibited the highest brightness and photostability under one-photon and two-photon excitation modes, respectively. All studied iRFPs exhibited efficient binding of the endogenous biliverdin chromophore in cultured neurons and in the mammalian brain and can be readily applied to neuroimaging.

PMID:
29017728
PMCID:
PMC5700256
DOI:
10.1016/j.bpj.2017.09.007
[Indexed for MEDLINE]
Free PMC Article

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