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Resonant out-of-phase fluorescence microscopy and remote imaging overcome spectral limitations
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Edité par CCSD ; Nature Publishing Group -
Author Correction: The Peer Review File associated with this Article was updated shortly after publication to redact from the authors’ point-by-point response a description of unpublished work describing how Speed OPIOM may in future be used to facilitate discrimination between FRET and direct excitation. DOI: 10.1038/s41467-017-02133-8. International audience. We present speed out-of-phase imaging after optical modulation (OPIOM), which exploits reversible photoswitchable fluorophores as fluorescent labels and combines optimized periodic illumination with phase-sensitive detection to specifically retrieve the label signal. Speed OPIOM can extract the fluorescence emission from a targeted label in the presence of spectrally interfering fluorophores and autofluorescence. Up to four fluorescent proteins exhibiting a similar green fluorescence have been distinguished in cells either sequentially or in parallel. Speed OPIOM is compatible with imaging biological processes in real time in live cells. Finally speed OPIOM is not limited to microscopy but is relevant for remote imaging as well, in particular, under ambient light. Thus, speed OPIOM has proved to enable fast and quantitative live microscopic and remote-multiplexed fluorescence imaging of biological samples while filtering out noise, interfering fluorophores, as well as ambient light.
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