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Light-induced cell damage in live-cell super-resolution microscopy

Super-resolution microscopy can unravel previously hidden details of cellular structures but requires high irradiation intensities to use the limited photon budget efficiently. Such high photon densities are likely to induce cellular damage in live-cell experiments. We applied single-molecule locali...

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Autores principales: Wäldchen, Sina, Lehmann, Julian, Klein, Teresa, van de Linde, Sebastian, Sauer, Markus
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4611486/
https://www.ncbi.nlm.nih.gov/pubmed/26481189
http://dx.doi.org/10.1038/srep15348
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author Wäldchen, Sina
Lehmann, Julian
Klein, Teresa
van de Linde, Sebastian
Sauer, Markus
author_facet Wäldchen, Sina
Lehmann, Julian
Klein, Teresa
van de Linde, Sebastian
Sauer, Markus
author_sort Wäldchen, Sina
collection PubMed
description Super-resolution microscopy can unravel previously hidden details of cellular structures but requires high irradiation intensities to use the limited photon budget efficiently. Such high photon densities are likely to induce cellular damage in live-cell experiments. We applied single-molecule localization microscopy conditions and tested the influence of irradiation intensity, illumination-mode, wavelength, light-dose, temperature and fluorescence labeling on the survival probability of different cell lines 20–24 hours after irradiation. In addition, we measured the microtubule growth speed after irradiation. The photo-sensitivity is dramatically increased at lower irradiation wavelength. We observed fixation, plasma membrane permeabilization and cytoskeleton destruction upon irradiation with shorter wavelengths. While cells stand light intensities of ~1 kW cm(−2) at 640 nm for several minutes, the maximum dose at 405 nm is only ~50 J cm(−2), emphasizing red fluorophores for live-cell localization microscopy. We also present strategies to minimize phototoxic factors and maximize the cells ability to cope with higher irradiation intensities.
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spelling pubmed-46114862015-11-02 Light-induced cell damage in live-cell super-resolution microscopy Wäldchen, Sina Lehmann, Julian Klein, Teresa van de Linde, Sebastian Sauer, Markus Sci Rep Article Super-resolution microscopy can unravel previously hidden details of cellular structures but requires high irradiation intensities to use the limited photon budget efficiently. Such high photon densities are likely to induce cellular damage in live-cell experiments. We applied single-molecule localization microscopy conditions and tested the influence of irradiation intensity, illumination-mode, wavelength, light-dose, temperature and fluorescence labeling on the survival probability of different cell lines 20–24 hours after irradiation. In addition, we measured the microtubule growth speed after irradiation. The photo-sensitivity is dramatically increased at lower irradiation wavelength. We observed fixation, plasma membrane permeabilization and cytoskeleton destruction upon irradiation with shorter wavelengths. While cells stand light intensities of ~1 kW cm(−2) at 640 nm for several minutes, the maximum dose at 405 nm is only ~50 J cm(−2), emphasizing red fluorophores for live-cell localization microscopy. We also present strategies to minimize phototoxic factors and maximize the cells ability to cope with higher irradiation intensities. Nature Publishing Group 2015-10-20 /pmc/articles/PMC4611486/ /pubmed/26481189 http://dx.doi.org/10.1038/srep15348 Text en Copyright © 2015, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Wäldchen, Sina
Lehmann, Julian
Klein, Teresa
van de Linde, Sebastian
Sauer, Markus
Light-induced cell damage in live-cell super-resolution microscopy
title Light-induced cell damage in live-cell super-resolution microscopy
title_full Light-induced cell damage in live-cell super-resolution microscopy
title_fullStr Light-induced cell damage in live-cell super-resolution microscopy
title_full_unstemmed Light-induced cell damage in live-cell super-resolution microscopy
title_short Light-induced cell damage in live-cell super-resolution microscopy
title_sort light-induced cell damage in live-cell super-resolution microscopy
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4611486/
https://www.ncbi.nlm.nih.gov/pubmed/26481189
http://dx.doi.org/10.1038/srep15348
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