Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM)

Abstract

We have developed a high-resolution fluorescence microscopy method based on high-accuracy localization of photoswitchable fluorophores. In each imaging cycle, only a fraction of the fluorophores were turned on, allowing their positions to be determined with nanometer accuracy. The fluorophore positions obtained from a series of imaging cycles were used to reconstruct the overall image. We demonstrated an imaging resolution of 20 nm. This technique can, in principle, reach molecular-scale resolution.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: STORM with photo-switchable fluorophores.
Figure 2: The high localization accuracy of individual switches during each switching cycle defines the intrinsic resolution of STORM.
Figure 3: STORM can resolve structures with sub-diffraction-limit resolution.

Similar content being viewed by others

References

  1. Pohl, D.W. & Courjon, D. Near Field Optics (Kluwer, Dordrecht, 1993).

    Book  Google Scholar 

  2. Zipfel, W.R., Williams, R.M. & Webb, W.W. Nat. Biotechnol. 21, 1368–1376 (2003).

    Article  Google Scholar 

  3. Hell, S.W. Nat. Biotechnol. 21, 1347–1355 (2003).

    Article  CAS  Google Scholar 

  4. Gustafsson, M.G.L. Proc. Natl. Acad. Sci. USA 102, 13081–13086 (2005).

    Article  CAS  Google Scholar 

  5. Hofmann, M., Eggeling, C., Jakobs, S. & Hell, S.W. Proc. Natl. Acad. Sci. USA 102, 17565–17569 (2005).

    Article  CAS  Google Scholar 

  6. Gelles, J., Schnapp, B.J. & Sheetz, M.P. Nature 331, 450–453 (1988).

    Article  CAS  Google Scholar 

  7. van Oijen, A.M., Kohler, J., Schmidt, J., Muller, M. & Brakenhoff, G.J. Chem. Phys. Lett. 292, 183–187 (1998).

    Article  CAS  Google Scholar 

  8. Thompson, R.E., Larson, D.R. & Webb, W.W. Biophys. J. 82, 2775–2783 (2002).

    Article  CAS  Google Scholar 

  9. Yildiz, A. et al. Science 300, 2061–2065 (2003).

    Article  CAS  Google Scholar 

  10. Gordon, M.P., Ha, T. & Selvin, P.R. Proc. Natl. Acad. Sci. USA 101, 6462–6465 (2004).

    Article  CAS  Google Scholar 

  11. Qu, X., Wu, D., Mets, L. & Scherer, N.F. Proc. Natl. Acad. Sci. USA 101, 11298–11303 (2004).

    Article  CAS  Google Scholar 

  12. Lidke, K.A., Rieger, B., Jovin, T.M. & Heintzmann, R. Opt. Express 13, 7052–7062 (2005).

    Article  Google Scholar 

  13. Ram, S., Ward, E.S. & Ober, R.J. Proc. Natl. Acad. Sci. USA 103, 4457–4462 (2006).

    Article  CAS  Google Scholar 

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

    Article  Google Scholar 

  15. Heilemann, M., Margeat, E., Kasper, R., Sauer, M. & Tinnefeld, P. J. Am. Chem. Soc. 127, 3801–3806 (2005).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work is supported by in part by the US National Institutes of Health, the Defense Advance Research Projects Agency and a Packard Science and Engineering Fellowship (to X.Z.). X.Z. is a Howard Hughes Medical Institute Investigator.

Author information

Authors and Affiliations

Authors

Contributions

M.J.R. and M.B. conceived the STORM imaging concept. M.J.R., DNA imaging and data analysis; M.B., RecA imaging and data acquisition. X.Z. supervised the project.

Corresponding author

Correspondence to Xiaowei Zhuang.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Switching of Cy3-Cy5–labeled antibody. (PDF 45 kb)

Supplementary Fig. 2

Localization accuracy of single switches before and after drift correction. (PDF 49 kb)

Supplementary Fig. 3

Fluorescence time trace of DNA labeled with multiple switches over many STORM imaging cycles. (PDF 78 kb)

Supplementary Methods (PDF 50 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Rust, M., Bates, M. & Zhuang, X. Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM). Nat Methods 3, 793–796 (2006). https://doi.org/10.1038/nmeth929

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth929

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing