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PLoS One. 2013 Dec 31;8(12):e85221. doi: 10.1371/journal.pone.0085221. eCollection 2013.

Screening fluorescent voltage indicators with spontaneously spiking HEK cells.

Author information

1
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, United States of America.
2
Department of Neurology, Yale School of Medicine, and Neurorehabilitation Research Center, Veterans Affairs Hospital, West Haven, Connecticut, United States of America.
3
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, United States of America ; Department of Physics, Harvard University, Cambridge, Massachusetts, United States of America.

Abstract

Development of improved fluorescent voltage indicators is a key challenge in neuroscience, but progress has been hampered by the low throughput of patch-clamp characterization. We introduce a line of non-fluorescent HEK cells that stably express NaV 1.3 and KIR 2.1 and generate spontaneous electrical action potentials. These cells enable rapid, electrode-free screening of speed and sensitivity of voltage sensitive dyes or fluorescent proteins on a standard fluorescence microscope. We screened a small library of mutants of archaerhodopsin 3 (Arch) in spiking HEK cells and identified two mutants with greater voltage-sensitivity than found in previously published Arch voltage indicators.

PMID:
24391999
PMCID:
PMC3877367
DOI:
10.1371/journal.pone.0085221
[Indexed for MEDLINE]
Free PMC Article

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