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

Title:
Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T-lymphocytes
Authors:
Thestrup, Thomas; Litzlbauer, Julia; Bartholomäus, Ingo; Mues, Marsilius; Russo, Luigi; Dana, Hod; Kovalchuk, Yuri; Liang, Yajie; Kalamakis, Georgios; Laukat, Yvonne; Becker, Stefan; Witte, Gregor; Geiger, Anselm; Allen, Taylor; Rome, Lawrence C.; Chen, Tsai-Wen; Kim, Douglas S.; Garaschuk, Olga; Griesinger, Christian; Griesbeck, Oliver
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.
Citation:
Thestrup, T., J. Litzlbauer, I. Bartholomäus, M. Mues, L. Russo, H. Dana, Y. Kovalchuk, Y. Liang, G. Kalamakis, Y. Laukat, S. Becker, G. Witte, A. Geiger, T. Allen, L.C. Rome, T.-W.Chen, D.S. Kim, O. Garaschuk, C. Griesinger, and O. Griesbeck. 2014. "Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T-lymphocytes." Nature Methods 11(2): 175-182.
Publisher:
Nature Publishing Group
DATE ISSUED:
2014
Department:
Biology
Type:
Article
PUBLISHED VERSION:
10.1038/nmeth.2773
PERMANENT LINK:
http://hdl.handle.net/11282/566831

Full metadata record

DC FieldValue Language
dc.contributor.authorThestrup, Thomasen
dc.contributor.authorLitzlbauer, Juliaen
dc.contributor.authorBartholomäus, Ingoen
dc.contributor.authorMues, Marsiliusen
dc.contributor.authorRusso, Luigien
dc.contributor.authorDana, Hoden
dc.contributor.authorKovalchuk, Yurien
dc.contributor.authorLiang, Yajieen
dc.contributor.authorKalamakis, Georgiosen
dc.contributor.authorLaukat, Yvonneen
dc.contributor.authorBecker, Stefanen
dc.contributor.authorWitte, Gregoren
dc.contributor.authorGeiger, Anselmen
dc.contributor.authorAllen, Tayloren
dc.contributor.authorRome, Lawrence C.en
dc.contributor.authorChen, Tsai-Wenen
dc.contributor.authorKim, Douglas S.en
dc.contributor.authorGaraschuk, Olgaen
dc.contributor.authorGriesinger, Christianen
dc.contributor.authorGriesbeck, Oliveren
dc.date.accessioned2015-08-13T10:35:05Zen
dc.date.available2015-08-13T10:35:05Zen
dc.date.issued2014en
dc.identifier.citationThestrup, T., J. Litzlbauer, I. Bartholomäus, M. Mues, L. Russo, H. Dana, Y. Kovalchuk, Y. Liang, G. Kalamakis, Y. Laukat, S. Becker, G. Witte, A. Geiger, T. Allen, L.C. Rome, T.-W.Chen, D.S. Kim, O. Garaschuk, C. Griesinger, and O. Griesbeck. 2014. "Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T-lymphocytes." Nature Methods 11(2): 175-182.en
dc.identifier.issn1548-7091en
dc.identifier.urihttp://hdl.handle.net/11282/566831en
dc.description.abstractThe 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.en
dc.language.isoen_USen
dc.publisherNature Publishing Groupen
dc.identifier.doi10.1038/nmeth.2773en
dc.subject.departmentBiologyen
dc.titleOptimized ratiometric calcium sensors for functional in vivo imaging of neurons and T-lymphocytesen
dc.typeArticleen
dc.identifier.journalNature Methodsen
dc.subject.keywordNeuroscienceen
dc.subject.keywordImaging the immune systemen
dc.subject.keywordFluorescent proteinsen
dc.subject.keywordFluorescence imagingen
dc.identifier.volume11en
dc.identifier.issue2en
dc.identifier.startpage175en
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