Bacterial cell wall nanoimaging by autoblinking microscopy

Abstract Spurious blinking fluorescent spots are often seen in bacteria during single-molecule localization microscopy experiments. Although this ‘autoblinking’ phenomenon is widespread, its origin remains unclear. In Deinococcus strains, we observed particularly strong autoblinking at the periphery...

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Autores principales:	Kevin Floc’h, Françoise Lacroix, Liliana Barbieri, Pascale Servant, Remi Galland, Corey Butler, Jean-Baptiste Sibarita, Dominique Bourgeois, Joanna Timmins
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Single-molecule Localization Microscopy (SMLM) PALM Imaging Point Accumulation For Imaging In Nanoscale Topography (PAINT) Photoactivated Localization Microscopy (PALM) Deinococcus Strains Medicine R Science Q
Acceso en línea:	https://doaj.org/article/d254383b03654346b1b45c560c87eb17
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spelling	oai:doaj.org-article:d254383b03654346b1b45c560c87eb172021-12-02T15:09:05ZBacterial cell wall nanoimaging by autoblinking microscopy10.1038/s41598-018-32335-z2045-2322https://doaj.org/article/d254383b03654346b1b45c560c87eb172018-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-32335-zhttps://doaj.org/toc/2045-2322Abstract Spurious blinking fluorescent spots are often seen in bacteria during single-molecule localization microscopy experiments. Although this ‘autoblinking’ phenomenon is widespread, its origin remains unclear. In Deinococcus strains, we observed particularly strong autoblinking at the periphery of the bacteria, facilitating its comprehensive characterization. A systematic evaluation of the contributions of different components of the sample environment to autoblinking levels and the in-depth analysis of the photophysical properties of autoblinking molecules indicate that the phenomenon results from transient binding of fluorophores originating mostly from the growth medium to the bacterial cell wall, which produces single-molecule fluorescence through a Point Accumulation for Imaging in Nanoscale Topography (PAINT) mechanism. Our data suggest that the autoblinking molecules preferentially bind to the plasma membrane of bacterial cells. Autoblinking microscopy was used to acquire nanoscale images of live, unlabeled D. radiodurans and could be combined with PALM imaging of PAmCherry-labeled bacteria in two-color experiments. Autoblinking-based super-resolved images provided insight into the formation of septa in dividing bacteria and revealed heterogeneities in the distribution and dynamics of autoblinking molecules within the cell wall.Kevin Floc’hFrançoise LacroixLiliana BarbieriPascale ServantRemi GallandCorey ButlerJean-Baptiste SibaritaDominique BourgeoisJoanna TimminsNature PortfolioarticleSingle-molecule Localization Microscopy (SMLM)PALM ImagingPoint Accumulation For Imaging In Nanoscale Topography (PAINT)Photoactivated Localization Microscopy (PALM)Deinococcus StrainsMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-12 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Single-molecule Localization Microscopy (SMLM) PALM Imaging Point Accumulation For Imaging In Nanoscale Topography (PAINT) Photoactivated Localization Microscopy (PALM) Deinococcus Strains Medicine R Science Q
spellingShingle	Single-molecule Localization Microscopy (SMLM) PALM Imaging Point Accumulation For Imaging In Nanoscale Topography (PAINT) Photoactivated Localization Microscopy (PALM) Deinococcus Strains Medicine R Science Q Kevin Floc’h Françoise Lacroix Liliana Barbieri Pascale Servant Remi Galland Corey Butler Jean-Baptiste Sibarita Dominique Bourgeois Joanna Timmins Bacterial cell wall nanoimaging by autoblinking microscopy
description	Abstract Spurious blinking fluorescent spots are often seen in bacteria during single-molecule localization microscopy experiments. Although this ‘autoblinking’ phenomenon is widespread, its origin remains unclear. In Deinococcus strains, we observed particularly strong autoblinking at the periphery of the bacteria, facilitating its comprehensive characterization. A systematic evaluation of the contributions of different components of the sample environment to autoblinking levels and the in-depth analysis of the photophysical properties of autoblinking molecules indicate that the phenomenon results from transient binding of fluorophores originating mostly from the growth medium to the bacterial cell wall, which produces single-molecule fluorescence through a Point Accumulation for Imaging in Nanoscale Topography (PAINT) mechanism. Our data suggest that the autoblinking molecules preferentially bind to the plasma membrane of bacterial cells. Autoblinking microscopy was used to acquire nanoscale images of live, unlabeled D. radiodurans and could be combined with PALM imaging of PAmCherry-labeled bacteria in two-color experiments. Autoblinking-based super-resolved images provided insight into the formation of septa in dividing bacteria and revealed heterogeneities in the distribution and dynamics of autoblinking molecules within the cell wall.
format	article
author	Kevin Floc’h Françoise Lacroix Liliana Barbieri Pascale Servant Remi Galland Corey Butler Jean-Baptiste Sibarita Dominique Bourgeois Joanna Timmins
author_facet	Kevin Floc’h Françoise Lacroix Liliana Barbieri Pascale Servant Remi Galland Corey Butler Jean-Baptiste Sibarita Dominique Bourgeois Joanna Timmins
author_sort	Kevin Floc’h
title	Bacterial cell wall nanoimaging by autoblinking microscopy
title_short	Bacterial cell wall nanoimaging by autoblinking microscopy
title_full	Bacterial cell wall nanoimaging by autoblinking microscopy
title_fullStr	Bacterial cell wall nanoimaging by autoblinking microscopy
title_full_unstemmed	Bacterial cell wall nanoimaging by autoblinking microscopy
title_sort	bacterial cell wall nanoimaging by autoblinking microscopy
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/d254383b03654346b1b45c560c87eb17
work_keys_str_mv	AT kevinfloch bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT francoiselacroix bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT lilianabarbieri bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT pascaleservant bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT remigalland bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT coreybutler bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT jeanbaptistesibarita bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT dominiquebourgeois bacterialcellwallnanoimagingbyautoblinkingmicroscopy AT joannatimmins bacterialcellwallnanoimagingbyautoblinkingmicroscopy
_version_	1718387968279838720

Bacterial cell wall nanoimaging by autoblinking microscopy

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