Skip to main content

Thank you for visiting 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.

Imaging cellular ultrastructures using expansion microscopy (U-ExM)


Determining the structure and composition of macromolecular assemblies is a major challenge in biology. Here we describe ultrastructure expansion microscopy (U-ExM), an extension of expansion microscopy that allows the visualization of preserved ultrastructures by optical microscopy. This method allows for near-native expansion of diverse structures in vitro and in cells; when combined with super-resolution microscopy, it unveiled details of ultrastructural organization, such as centriolar chirality, that could otherwise be observed only by electron microscopy.

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

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type



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

Fig. 1: Centriole expansion with U-ExM.
Fig. 2: U-ExM reaches dSTORM precision limits.
Fig. 3: U-ExM applied to human cells.

Data availability

The data that support the findings of this study are available from the corresponding authors upon request.


  1. Koster, A. J. & Klumperman, J. Nat. Rev. Mol. Cell Biol. 4, SS6–SS10 (2003).

    Google Scholar 

  2. Sahl, S. J., Hell, S. W. & Jakobs, S. Nat. Rev. Mol. Cell Biol. 18, 685–701 (2017).

    Article  CAS  Google Scholar 

  3. Chen, F., Tillberg, P. W. & Boyden, E. S. Science 347, 543–548 (2015).

    Article  CAS  Google Scholar 

  4. Chozinski, T. J. et al. Nat. Methods 13, 485–488 (2016).

    Article  CAS  Google Scholar 

  5. Tillberg, P. W. et al. Nat. Biotechnol. 34, 987–992 (2016).

    Article  CAS  Google Scholar 

  6. Ku, T. et al. Nat. Biotechnol. 34, 973–981 (2016).

    Article  CAS  Google Scholar 

  7. Hamel, V. et al. Curr. Biol. 27, 2486–2498 (2017).

    Article  CAS  Google Scholar 

  8. Klena, N. et al. J. Vis. Exp. 2018, e58109 (2018).

    Google Scholar 

  9. Heilemann, M. et al. Angew. Chem. Int. Ed. Engl. 47, 6172–6176 (2008).

    Article  CAS  Google Scholar 

  10. Fortun, D. et al. IEEE Trans. Med. Imaging 37, 1235–1246 (2018).

    Article  Google Scholar 

  11. Heine, J. et al. Proc. Natl Acad. Sci. USA 114, 9797–9802 (2017).

  12. Pigino, G. et al. J. Cell. Biol. 195, 673–687 (2011).

    Article  CAS  Google Scholar 

  13. Lechtreck, K. F. & Geimer, S. Cell Motil. Cytoskeleton 47, 219–235 (2000).

    Article  CAS  Google Scholar 

  14. Kubo, T., Yanagisawa, H. A., Yagi, T., Hirono, M. & Kamiya, R. Curr. Biol. 20, 441–445 (2010).

    Article  CAS  Google Scholar 

  15. Kubo, T. & Oda, T. Mol. Biol. Cell. 28, 2260–2266 (2017).

    Article  CAS  Google Scholar 

  16. Suryavanshi, S. et al. Curr. Biol. 20, 435–440 (2010).

    Article  CAS  Google Scholar 

  17. Gao, M. et al. ACS Nano 12, 4178–4185 (2018).

    Article  CAS  Google Scholar 

  18. Halpern, A. R., Alas, G. C. M., Chozinski, T. J., Paredez, A. R. & Vaughan, J. C. ACS Nano 11, 12677–12686 (2017).

    Article  CAS  Google Scholar 

  19. Yang, T. T. et al. Nat. Commun. 9, 2023 (2018).

    Article  Google Scholar 

  20. Borlinghaus, R. T. & Kappel, C. Nat. Methods 13, i–iii (2016).

  21. De Luca, G. M. R. et al. J. Microsc. 266, 166–177 (2017).

  22. Göttfert, F. et al. Proc. Natl Acad. Sci. USA 114, 2125–2130 (2017).

  23. Ovesný, M., Křížek, P., Borkovec, J., Švindrych, Z. & Hagen, G. M. Bioinformatics 30, 2389–2390 (2014).

    Article  Google Scholar 

  24. Wolter, S. et al. Nat. Methods 9, 1040–1041 (2012).

    Article  CAS  Google Scholar 

  25. Thévenaz, P., Ruttimann, U. E. & Unser, M. IEEE Trans. Image Process. 7, 27–41 (1998).

    Article  Google Scholar 

  26. Unser, M., Soubies, E., Soulez, F., McCann, M. & Donati, L. GlobalBioIm: a unifying computational framework for solving inverse problems. OSA Technical Digest (2017).

  27. Schindelin, J. et al. Nat. Methods 9, 676–682 (2012).

    Article  CAS  Google Scholar 

  28. Schneider, C. A., Rasband, W. S. & Eliceiri, K. W. Nat. Methods 9, 671–675 (2012).

    Article  CAS  Google Scholar 

  29. Royer, L. A. et al. Nat. Methods 12, 480–481 (2015).

    Article  CAS  Google Scholar 

Download references


We thank N. Klena for critical reading of the manuscript. We thank the BioImaging Center (University of Geneva) for help in image acquisition. We thank the Martinou lab and especially S. Zaganelli for helpful discussions and sharing of mitochondrial reagents. Human U2OS cells were a gift from E. Nigg (Biozentrum, University of Basel, Basel, Switzerland). D.G. and M.S.-C. are supported by the European Research Council (ERC; StG 715289 (ACCENT)). P.G., V.H., and M.L.G. are supported by the Swiss National Science Foundation (SNSF; PP00P3_157517). F.U.Z. and M.S. acknowledge support by the Deutsche Forschungsgemeinschaft (DFG) within the Collaborative Research Center 166 ReceptorLight (projects A04 and B04). M.U. is supported by the ERC (GA No. 692726 GlobalBioIm).

Author information

Authors and Affiliations



D.G., F.U.Z., M.S., V.H., and P.G. conceived and designed the project. M.S., V.H., and P.G. supervised the project. D.G. and F.U.Z. performed all ExM experiments. D.G. performed all U-ExM experiments with the help of S.B., as well as the data analysis. F.U.Z. performed the dSTORM imaging and the experiment and analysis involving clathrin-coated pits. J.H., J.-G.S., and M.R. performed and analyzed the STED imaging. D.F. and M.U. performed the 3D averaging. M.S.-C. initiated the U-ExM project. M.L.G. performed the plot profile of the polar transform showing the ninefold symmetry, as well as the r.m.s. calculation. E.S.B. helped in setting up ExM. All authors wrote and revised the final manuscript.

Corresponding authors

Correspondence to Markus Sauer, Virginie Hamel or Paul Guichard.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary text and figures

Supplementary Figures 1–19

Reporting Summary

Supplementary Video 1

U-ExM of isolated Chlamydomonas centrioles. Confocal (HyVolution) stack of immunolabeled Chlamydomonas centrioles in U-ExM. Magenta corresponds to α-tubulin, and green to PolyE. Scale bar, 1 µm.

Supplementary Video 2

3D rendering of isolated Chlamydomonas centrioles treated by U-ExM. 3D rendering of the confocal stack from Supplementary Video 1. Note the polyglutamylation signal (PolyE; green) along the microtubule triplets of the mature centrioles. Scale bar, 1 µm.

Supplementary Video 3

DyMIN imaging of U-ExM isolated Chlamydomonas centrioles. Stack of immunolabeled Chlamydomonas centrioles treated by U-ExM and imaged with DyMIN. Magenta corresponds to α-tubulin, and green to PolyE. Scale bar, 1 µm.

Supplementary Video 4

Supplementary Video 4U-ExM of Chlamydomonas cell. Confocal (HyVolution) stack of an immunolabeled Chlamydomonas cell treated by U-ExM. Magenta corresponds to α-tubulin, and green to PolyE.

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Gambarotto, D., Zwettler, F.U., Le Guennec, M. et al. Imaging cellular ultrastructures using expansion microscopy (U-ExM). Nat Methods 16, 71–74 (2019).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


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