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Ultrabright photoactivatable fluorophores created by reductive caging

Nature Methods volume 9, pages 1181–1184 (2012)Cite this article

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Abstract

Sub–diffraction-limit imaging can be achieved by sequential localization of photoactivatable fluorophores, for which the image resolution depends on the number of photons detected per localization. We report a strategy for fluorophore caging that creates photoactivatable probes with high photon yields. Upon photoactivation, these probes can provide 104−106 photons per localization and allow imaging of fixed samples with resolutions of several nanometers. This strategy can be applied to many fluorophores across the visible spectrum.

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Figure 1: Bright photoactivatable dyes created by reductive caging.
Figure 2: STORM images of microtubules in cells stained by indirect immunofluorescence with blue, yellow and red dyes.
Figure 3: STORM image and transverse profile characterization of microtubules polymerized and labeled in vitro with Cy3B.

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Acknowledgements

We thank T. Mitchison, M. Ericsson and W. Wang for help during this project. This work is supported in part by the US National Institutes of Health and a Collaborative Innovation Award from the Howard Hughes Medical Institute (to X.Z.). J.C.V. is supported in part by a Burroughs-Wellcome Career Award at the Scientific Interface. X.Z. is a Howard Hughes Medical Institute investigator.

Author information

Authors and Affiliations

  1. Howard Hughes Medical Institute, Harvard University, Cambridge, Massachusetts, USA

    Joshua C Vaughan, Shu Jia & Xiaowei Zhuang

  2. Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, USA

    Joshua C Vaughan, Shu Jia & Xiaowei Zhuang

  3. Department of Physics, Harvard University, Cambridge, Massachusetts, USA

    Xiaowei Zhuang

Authors
  1. Joshua C Vaughan
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  2. Shu Jia
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Contributions

J.C.V. is primarily responsible for experimental design. J.C.V. and S.J. performed experiments and analysis. X.Z. supervised and guided the project. J.C.V. and X.Z. wrote the manuscript.

Corresponding author

Correspondence to Xiaowei Zhuang.

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The authors declare no competing financial interests.

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Supplementary Figures 1–9, Supplementary Note and Supplementary Protocol (PDF 9669 kb)

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Vaughan, J., Jia, S. & Zhuang, X. Ultrabright photoactivatable fluorophores created by reductive caging. Nat Methods 9, 1181–1184 (2012). https://doi.org/10.1038/nmeth.2214

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