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  Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes

Thestrup, T., Litzlbauer, J., Bartholomäus, I., Mues, M., Russo, L., Dana, H., et al. (2014). Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes. NATURE METHODS, 11(2), 175-182. doi:10.1038/NMETH.2773.

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 Creators:
Thestrup, Thomas1, Author              
Litzlbauer, Julia1, Author              
Bartholomäus, Ingo2, Author              
Mues, Marsilius2, Author              
Russo, Luigi3, Author
Dana, Hod3, Author
Kovalchuk, Yuri3, Author
Liang, Yajie3, Author
Kalamakis, Georgios3, Author
Laukat, Yvonne3, Author
Becker, Stefan3, Author
Witte, Gregor3, Author
Geiger, Anselm1, Author              
Allen, Taylor3, Author
Rome, Lawrence C.3, Author
Chen, Tsai-Wen3, Author
Kim, Douglas S.3, Author
Garaschuk, Olga3, Author
Griesinger, Christian3, Author
Griesbeck, Oliver1, Author              
Affiliations:
1Research Group: Cellular Dynamics / Griesbeck, MPI of Neurobiology, Max Planck Society, ou_1113560              
2Emeritus Group: Neuroimmunology / Wekerle, MPI of Neurobiology, Max Planck Society, ou_1113547              
3external, ou_persistent22              

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Free keywords: X-RAY SOLUTION; GREEN FLUORESCENT PROTEIN; ENCODED CA2+ INDICATORS; SMALL-ANGLE SCATTERING; MOUSE VISUAL-CORTEX; NEURAL ACTIVITY; LARGER PROTEINS; TROPONIN-C; MACROMOLECULAR STRUCTURES; NMR
 Abstract: The quality of genetically encoded calcium indicators (GECIs) has improved dramatically in recent years, but high-performing ratiometric indicators are still rare. Here we describe a series of fluorescence resonance energy transfer (FRET)-based calcium biosensors with a reduced number of calcium binding sites per sensor. These 'Twitch' sensors are based on the C-terminal domain of Opsanus troponin C. Their FRET responses were optimized by a large-scale functional screen in bacterial colonies, refined by a secondary screen in rat hippocampal neuron cultures. We tested the in vivo performance of the most sensitive variants in the brain and lymph nodes of mice. The sensitivity of the Twitch sensors matched that of synthetic calcium dyes and allowed visualization of tonic action potential firing in neurons and high resolution functional tracking of T lymphocytes. Given their ratiometric readout, their brightness, large dynamic range and linear response properties, Twitch sensors represent versatile tools for neuroscience and immunology.

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Language(s): eng - English
 Dates: 2014-02
 Publication Status: Published in print
 Pages: 14
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: ISI: 000331141600020
DOI: 10.1038/NMETH.2773
 Degree: -

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Title: NATURE METHODS
Source Genre: Journal
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Publ. Info: MACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLAND : NATURE PUBLISHING GROUP
Pages: - Volume / Issue: 11 (2) Sequence Number: - Start / End Page: 175 - 182 Identifier: ISSN: 1548-7091