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Screening fluorescent voltage probes with spontaneously spiking HEK cells (779.5)
Author(s) -
Park Jeehae,
Werley Christopher,
Cohen Adam
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.779.5
Subject(s) - hek 293 cells , fluorescence , voltage clamp , patch clamp , fluorescence microscope , biophysics , mutant , green fluorescent protein , electrophysiology , chemistry , microbiology and biotechnology , cell culture , biology , neuroscience , biochemistry , membrane potential , physics , genetics , optics , gene
A major difficulty for imaging brain activity comes from the lack of voltage‐sensitive probes that have fast, sensitive and bright characteristics. Unlike other probe development challenges, screening fluorescent voltage probes 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.