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)

Nature Methods volume 3, pages793–796(2006)Cite this article

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.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

Rent or Buy

All prices are NET prices.

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.

References

  1. 1

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

    Google Scholar 

  2. 2

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

    Article  Google Scholar 

  3. 3

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

    CAS  Article  Google Scholar 

  4. 4

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

    CAS  Article  Google Scholar 

  5. 5

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

    CAS  Article  Google Scholar 

  6. 6

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

    CAS  Article  Google Scholar 

  7. 7

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

    CAS  Article  Google Scholar 

  8. 8

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

    CAS  Article  Google Scholar 

  9. 9

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

    CAS  Article  Google Scholar 

  10. 10

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

    CAS  Article  Google Scholar 

  11. 11

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

    CAS  Article  Google Scholar 

  12. 12

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

    Article  Google Scholar 

  13. 13

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

    CAS  Article  Google Scholar 

  14. 14

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

    Article  Google Scholar 

  15. 15

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

    CAS  Article  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

Author notes

  1. Michael J Rust and Mark Bates: These authors contributed equally to this work.

Affiliations

  1. Department of Physics, Harvard University, 12 Oxford Street, Cambridge, 02138, Massachusetts, USA

    Michael J Rust & Xiaowei Zhuang

  2. Division of Engineering and Applied Sciences, Harvard University, 12 Oxford Street, Cambridge, 02138, Massachusetts, USA

    Mark Bates

  3. Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, 02138, Massachusetts, USA

    Xiaowei Zhuang

  4. Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, 02138, Massachusetts, USA

    Xiaowei Zhuang

Authors
  1. Michael J Rust
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mark Bates
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xiaowei Zhuang
    View author publications

    You can also search for this author in PubMed Google Scholar

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

Further reading

Search

Advanced 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