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Two-color far-field fluorescence nanoscopy based on photoswitchable emitters

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Abstract

We demonstrate two-color far-field fluorescence microscopy with nanoscale spatial resolution based on the photoswitching of individual fluorescent markers. By enabling, recording, and disabling the emission of the reversibly switchable fluorescent protein rsFastLime and of the organic fluorophore cyanine5, we recorded two-color nanoscale images inside whole cells. The position of individual emitters was determined with a typical accuracy of 20 nm, which largely constitutes the lateral resolution of the system. Photoswitching in two-color colocalization experiments represents a major step towards the application of far-field fluorescence nanoscopy to the study of (biological) samples on the macromolecular level.

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References

  1. E. Abbe, Arch. Mikrosk. Anat. 9, 413 (1873)

    Google Scholar 

  2. S.W. Hell, Science 316, 1153 (2007)

    Article  ADS  Google Scholar 

  3. S.W. Hell, Opt. Commun. 106, 19 (1994)

    Article  ADS  Google Scholar 

  4. S.W. Hell, J. Wichmann, Opt. Lett. 19, 780 (1994)

    ADS  Google Scholar 

  5. S.W. Hell, S. Jakobs, L. Kastrup, Appl. Phys. A 77, 859 (2003)

    Article  ADS  Google Scholar 

  6. M. Hofmann, C. Eggeling, S. Jakobs, S.W. Hell, Proc. Nat. Acad. Sci. 102, 17565 (2005)

    Article  ADS  Google Scholar 

  7. E. Betzig, G.H. Patterson, R. Sougrat, O.W. Lindwasser, S. Olenych, J.S. Bonifacino, M.W. Davidson, J. Lippincott-Schwartz, H.F. Hess, Science 313, 1642 (2006)

    Article  ADS  Google Scholar 

  8. S.T. Hess, T.P.K. Girirajan, M.D. Mason, Biophys. J. 91, 4258 (2006)

    Article  ADS  Google Scholar 

  9. M.J. Rust, M. Bates, X. Zhuang, Nature Methods 3, 793 (2006)

    Google Scholar 

  10. E. Betzig, Opt. Lett. 20, 237 (1995)

    ADS  Google Scholar 

  11. G.J. Schütz, H. Schindler, T. Schmidt, Biophys. J. 73, 1073 (1997)

    Article  Google Scholar 

  12. T.D. Lacoste, X. Michalet, F. Pinaud, D.S. Chemla, A.P. Alivisatos, S. Weiss, Proc. Nat. Acad. Sci. 97, 9461 (2000)

    Article  ADS  Google Scholar 

  13. R.E. Thompson, D.R. Larson, W.W. Webb, Biophys. J. 82, 2775 (2002)

    Google Scholar 

  14. S.W. Hell, J. Soukka, P.E. Hänninen, Bioimaging 3, 65 (1995)

    Article  Google Scholar 

  15. C. Geisler, A. Schönle, C. von Middendorff, H. Bock, C. Eggeling, A. Egner, S.W. Hell, Appl. Phys. A 88, 223 (2007)

    Article  ADS  Google Scholar 

  16. E.A. Jares-Erijman, T.M. Jovin, Nature Biotechnol. 21, 1387 (2003)

    Google Scholar 

  17. G. Donnert, J. Keller, C.A. Wurm, S.O. Rizzoli, V. Westphal, A. Schönle, R. Jahn, S. Jakobs, C. Eggeling, S.W. Hell, Biophys. J. 92, L67 (2007)

    Article  Google Scholar 

  18. A.C. Stiel, S. Trowitzsch, G. Weber, M. Andresen, C. Eggeling, S.W. Hell, S. Jakobs, M.C. Wahl, Biochem. J. 402, 35 (2007)

    Article  Google Scholar 

  19. R. Ando, H. Mizuno, A. Miyawaki, Science 306, 1370 (2004)

    Article  ADS  Google Scholar 

  20. S. Habuchi, R. Ando, P. Dedecker, W. Verheijen, H. Mizuno, A. Miyawaki, J. Hofkens, Proc. Nat. Acad. Sci. 102, 9511 (2005)

    Article  ADS  Google Scholar 

  21. M. Bates, T.R. Blosser, X. Zhuang, Phys. Rev. Lett. 94, 108101 (2005)

    Article  ADS  Google Scholar 

  22. M. Heilemann, E. Margeat, R. Kasper, M. Sauer, P. Tinnefeld, J. Am. Chem. Soc. 127, 3801 (2005)

    Article  Google Scholar 

  23. R. Zondervan, F. Kulzer, M.A. Kolchenko, M. Orrit, J. Phys. Chem. A 108, 1657 (2004)

    Article  Google Scholar 

  24. Z. Huang, D. Ji, A. Xia, Colloid Surf. A 257258, 203 (2005)

    Google Scholar 

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Correspondence to S.W. Hell or C. Eggeling.

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33.50.Dq; 42.30.-d; 87.64.-t

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Bock, H., Geisler, C., Wurm, C. et al. Two-color far-field fluorescence nanoscopy based on photoswitchable emitters. Appl. Phys. B 88, 161–165 (2007). https://doi.org/10.1007/s00340-007-2729-0

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  • DOI: https://doi.org/10.1007/s00340-007-2729-0

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