PMID- 24390440
OWN - NLM
STAT- MEDLINE
DCOM- 20140325
LR  - 20220309
IS  - 1548-7105 (Electronic)
IS  - 1548-7091 (Linking)
VI  - 11
IP  - 2
DP  - 2014 Feb
TI  - Optimized ratiometric calcium sensors for functional in vivo imaging of neurons 
      and T lymphocytes.
PG  - 175-82
LID - 10.1038/nmeth.2773 [doi]
AB  - 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.
FAU - Thestrup, Thomas
AU  - Thestrup T
AD  - 1] Max Planck Institute of Neurobiology, Martinsried, Germany. [2] Janelia Farm 
      Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, USA.
FAU - Litzlbauer, Julia
AU  - Litzlbauer J
AD  - Max Planck Institute of Neurobiology, Martinsried, Germany.
FAU - Bartholomaus, Ingo
AU  - Bartholomaus I
AD  - Max Planck Institute of Neurobiology, Martinsried, Germany.
FAU - Mues, Marsilius
AU  - Mues M
AD  - Max Planck Institute of Neurobiology, Martinsried, Germany.
FAU - Russo, Luigi
AU  - Russo L
AD  - Max Planck Institute for Biophysical Chemistry, NMR-based Structural Biology, 
      Gottingen, Germany.
FAU - Dana, Hod
AU  - Dana H
AD  - Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, 
      USA.
FAU - Kovalchuk, Yuri
AU  - Kovalchuk Y
AD  - Institute of Physiology II, University of Tubingen, Tubingen, Germany.
FAU - Liang, Yajie
AU  - Liang Y
AUID- ORCID: 0000000247984882
AD  - Institute of Physiology II, University of Tubingen, Tubingen, Germany.
FAU - Kalamakis, Georgios
AU  - Kalamakis G
AD  - Institute of Physiology II, University of Tubingen, Tubingen, Germany.
FAU - Laukat, Yvonne
AU  - Laukat Y
AD  - Max Planck Institute for Biophysical Chemistry, NMR-based Structural Biology, 
      Gottingen, Germany.
FAU - Becker, Stefan
AU  - Becker S
AD  - Max Planck Institute for Biophysical Chemistry, NMR-based Structural Biology, 
      Gottingen, Germany.
FAU - Witte, Gregor
AU  - Witte G
AUID- ORCID: 0000000303130962
AD  - Gene Center, Department of Biochemistry, University of Munich, Munich, Germany.
FAU - Geiger, Anselm
AU  - Geiger A
AD  - Max Planck Institute of Neurobiology, Martinsried, Germany.
FAU - Allen, Taylor
AU  - Allen T
AUID- ORCID: 000000033018187X
AD  - Oberlin College, Oberlin, Ohio, USA.
FAU - Rome, Lawrence C
AU  - Rome LC
AD  - 1] Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania, 
      USA. [2] The Whitman Center, Marine Biological Lab, Woods Hole, Massachusetts, 
      USA.
FAU - Chen, Tsai-Wen
AU  - Chen TW
AD  - Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, 
      USA.
FAU - Kim, Douglas S
AU  - Kim DS
AD  - Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, 
      USA.
FAU - Garaschuk, Olga
AU  - Garaschuk O
AD  - Institute of Physiology II, University of Tubingen, Tubingen, Germany.
FAU - Griesinger, Christian
AU  - Griesinger C
AD  - Max Planck Institute for Biophysical Chemistry, NMR-based Structural Biology, 
      Gottingen, Germany.
FAU - Griesbeck, Oliver
AU  - Griesbeck O
AD  - 1] Max Planck Institute of Neurobiology, Martinsried, Germany. [2] Janelia Farm 
      Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, USA.
LA  - eng
SI  - GENBANK/KF843821
SI  - PDB/2M97
GR  - Howard Hughes Medical Institute/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
DEP - 20140105
PL  - United States
TA  - Nat Methods
JT  - Nature methods
JID - 101215604
RN  - 0 (Fluorescent Dyes)
RN  - 0 (Luminescent Proteins)
RN  - 0 (Troponin C)
RN  - SY7Q814VUP (Calcium)
SB  - IM
CIN - Nat Methods. 2014 Feb;11(2):139-40. PMID: 24481218
MH  - Animals
MH  - Animals, Newborn
MH  - Biosensing Techniques/*methods
MH  - Calcium/*metabolism
MH  - Fluorescence Resonance Energy Transfer
MH  - Fluorescent Dyes
MH  - HEK293 Cells
MH  - Hippocampus/*metabolism
MH  - Humans
MH  - Image Processing, Computer-Assisted
MH  - Luminescent Proteins/*metabolism
MH  - Lymphocyte Activation
MH  - Magnetic Resonance Spectroscopy
MH  - Mice
MH  - Mice, Inbred C57BL
MH  - Microscopy, Fluorescence
MH  - Molecular Sequence Data
MH  - Neurons/cytology/*metabolism
MH  - Rats
MH  - T-Lymphocytes/cytology/*metabolism
MH  - Troponin C/*metabolism
EDAT- 2014/01/07 06:00
MHDA- 2014/03/26 06:00
CRDT- 2014/01/07 06:00
PHST- 2013/06/15 00:00 [received]
PHST- 2013/11/29 00:00 [accepted]
PHST- 2014/01/07 06:00 [entrez]
PHST- 2014/01/07 06:00 [pubmed]
PHST- 2014/03/26 06:00 [medline]
AID - nmeth.2773 [pii]
AID - 10.1038/nmeth.2773 [doi]
PST - ppublish
SO  - Nat Methods. 2014 Feb;11(2):175-82. doi: 10.1038/nmeth.2773. Epub 2014 Jan 5.