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Assessing phototoxicity in live fluorescence imaging

Abstract

Are the answers to biological questions obtained via live fluorescence microscopy substantially affected by phototoxicity? Although a single set of standards for assessing phototoxicity cannot exist owing to the breadth of samples and experimental questions associated with biological imaging, we need quantitative, practical assessments and reporting standards to ensure that imaging has a minimal impact on observed biological processes and sample health. Here we discuss the problem of phototoxicity in biology and suggest guidelines to improve its reporting and assessment.

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Figure 1: The effect of low levels of blue light on cell proliferation.
Figure 2: The four main considerations for live imaging.
Figure 3: Identifying species-specific criteria for the assessment of phototoxicity.

References

  1. Magidson, V. & Khodjakov, A. Methods Cell Biol. 114, 545–560 (2013).

    Article  Google Scholar 

  2. Stelzer, E.H.K. Nat. Methods 12, 23–26 (2015).

    Article  CAS  Google Scholar 

  3. Jemielita, M., Taormina, M.J., Delaurier, A., Kimmel, C.B. & Parthasarathy, R. J. Biophotonics 6, 920–928 (2013).

    Article  CAS  Google Scholar 

  4. Tinevez, J.-Y. et al. Methods Enzymol. 506, 291–309 (2012).

    Article  CAS  Google Scholar 

  5. Waters, J.C. & Wittmann, T. (eds.) Quantitative Imaging in Cell Biology. (Academic Press, 2014).

    Google Scholar 

  6. Goldman, R.D., Swedlow, J.R. & Spector, D.L. (eds.) Live Cell Imaging: A Laboratory Manual 2nd ed. (Cold Spring Harbor Laboratory Press, 2009).

    Google Scholar 

  7. Daddysman, M.K., Tycon, M.A. & Fecko, C.J. Methods Mol. Biol. 1148, 1–17 (2014).

    Article  CAS  Google Scholar 

  8. Cole, R. Cell Adh. Migr. 8, 452–459 (2014).

    Article  Google Scholar 

  9. Cranfill, P.J. et al. Nat. Methods 13, 557–562 (2016).

    Article  CAS  Google Scholar 

  10. Donnert, G., Eggeling, C. & Hell, S.W. Nat. Methods 4, 81–86 (2007).

    Article  CAS  Google Scholar 

  11. Murray, J.M., Appleton, P.L., Swedlow, J.R. & Waters, J.C. J. Microsc. 228, 390–405 (2007).

    Article  Google Scholar 

  12. Murray, J.M. An icosahedral virus as a fluorescent calibration standard: methods for counting protein molecules in cells by fluorescence microscopy. Preprint at http://biorxiv.org/content/early/2017/02/18/088617 (2016).

  13. Spielmann, H. et al. Altern. Lab. Anim. 28, 777–814 (2000).

    CAS  PubMed  Google Scholar 

  14. Bianchi, J.I., Stockert, J.C., Buzzi, L.I., Blázquez-Castro, A. & Simonetta, S.H. PLoS One 10, e0128898 (2015).

    Article  Google Scholar 

  15. AlGhamdi, K.M., Kumar, A. & Moussa, N.A. Lasers Med. Sci. 27, 237–249 (2012).

    Article  Google Scholar 

  16. Harnett, M.T., Makara, J.K., Spruston, N., Kath, W.L. & Magee, J.C. Nature 491, 599–602 (2012).

    Article  CAS  Google Scholar 

  17. Galli, R. et al. PLoS One 9, e110295 (2014).

    Article  Google Scholar 

  18. Logg, K., Bodvard, K., Blomberg, A. & Käll, M. FEMS Yeast Res. 9, 875–884 (2009).

    Article  CAS  Google Scholar 

  19. Knoll, S.G., Ahmed, W.W. & Saif, T.A. Sci. Rep. 5, 18513 (2015).

    Article  CAS  Google Scholar 

  20. Roehlecke, C., Schaller, A., Knels, L. & Funk, R.H.W. Mol. Vis. 15, 1929–1938 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. McDonald, A., Harris, J., Macmillan, D., Dempster, J. & McConnell, G. Biomed. Opt. Express 3, 1266–1273 (2012).

    Article  CAS  Google Scholar 

  22. Wäldchen, S., Lehmann, J., Klein, T., van de Linde, S. & Sauer, M. Sci. Rep. 5, 15348 (2015).

    Article  Google Scholar 

  23. Tomer, R., Khairy, K., Amat, F. & Keller, P.J. Nat. Methods 9, 755–763 (2012).

    Article  CAS  Google Scholar 

  24. Wu, Y. et al. Proc. Natl. Acad. Sci. USA 108, 17708–17713 (2011).

    Article  CAS  Google Scholar 

  25. Icha, J., Kunath, C., Rocha-Martins, M. & Norden, C. J. Cell Biol. 215, 259–275 (2016).

    Article  CAS  Google Scholar 

  26. Strnad, P. et al. Nat. Methods 13, 139–142 (2016).

    Article  CAS  Google Scholar 

  27. Strobl, F. & Stelzer, E.H.K. Development 141, 2331–2338 (2014).

    Article  CAS  Google Scholar 

  28. Carlton, P.M. et al. Proc. Natl. Acad. Sci. USA 107, 16016–16022 (2010).

    Article  CAS  Google Scholar 

  29. König, K. in Methods in Cellular Imaging (ed. Periasamy, A.) 236–251 (Oxford University Press, 2001).

    Book  Google Scholar 

  30. Hoebe, R.A. et al. Nat. Biotechnol. 25, 249–253 (2007).

    Article  CAS  Google Scholar 

  31. Kuznetsova, D.S. et al. J. Biophotonics 8, 952–960 (2015).

    Article  CAS  Google Scholar 

  32. Icha, J. et al. J. Vis. Exp. 110, e53966 (2016).

    Google Scholar 

  33. Dobrucki, J.W., Feret, D. & Noatynska, A. Biophys. J. 93, 1778–1786 (2007).

    Article  CAS  Google Scholar 

  34. Wu, Y. et al. Sci. Rep. 5, 14766 (2015).

    Article  CAS  Google Scholar 

  35. Boudreau, C. et al. Sci. Rep. 6, 30892 (2016).

    Article  CAS  Google Scholar 

  36. Chen, B.-C. et al. Science 346, 1257998 (2014).

    Article  Google Scholar 

  37. Ji, N., Magee, J.C. & Betzig, E. Nat. Methods 5, 197–202 (2008).

    Article  CAS  Google Scholar 

  38. US Environmental Protection Agency. Guidelines for Exposure Assessment. Report No. EPA/600/Z-92/001 (US EPA, 1992).

Download references

Acknowledgements

The authors thank all the delegates of LSFM2016 who attended and contributed to an exciting roundtable discussion. LSFM2016 was supported by the Company of Biologists (EA1170). P.P.L. thanks R. Heintzmann, S. Fraser, J. Huisken, and J.A. Laissue for discussions on phototoxicity. This work was supported by the Royal Society (IF150018 to P.P.L.) and by King Abdulaziz University, Rabigh Campus, Jeddah, Saudi Arabia (scholarship to R.A.A.).

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Correspondence to P Philippe Laissue.

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Laissue, P., Alghamdi, R., Tomancak, P. et al. Assessing phototoxicity in live fluorescence imaging. Nat Methods 14, 657–661 (2017). https://doi.org/10.1038/nmeth.4344

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