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.

GraFix: sample preparation for single-particle electron cryomicroscopy


We developed a method, named GraFix, that considerably improves sample quality for structure determination by single-particle electron cryomicroscopy (cryo-EM). GraFix uses a glycerol gradient centrifugation step in which the complexes are centrifuged into an increasing concentration of a chemical fixation reagent to prevent aggregation and to stabilize individual macromolecules. The method can be used to prepare samples for negative-stain, cryo-negative-stain and, particularly, unstained cryo-EM.

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

Figure 1: Schematics of the GraFix setup.
Figure 2: Effect of GraFix-based sample preparation on B complex spliceosomes and the 70S ribosome.
Figure 3: Improved structure of GraFix-prepared APC/C.


  1. Gavin, A.C. et al. Nature 440, 631–636 (2006).

    Article  CAS  Google Scholar 

  2. van Heel, M. et al. Q. Rev. Biophys. 33, 307–369 (2000).

    Article  CAS  Google Scholar 

  3. Chiu, W. et al. Structure 13, 363–372 (2005).

    Article  CAS  Google Scholar 

  4. Golas, M.M. et al. Science 300, 980–984 (2003).

    Article  CAS  Google Scholar 

  5. Halic, M. et al. Science 312, 745–747 (2006).

    Article  CAS  Google Scholar 

  6. Boehringer, D. et al. Nat. Struct. Mol. Biol. 11, 463–468 (2004).

    Article  CAS  Google Scholar 

  7. Dube, P. et al. Mol. Cell 20, 867–879 (2005).

    Article  CAS  Google Scholar 

  8. Migneault, I. et al. Biotechniques 37, 790–802 (2004).

    Article  CAS  Google Scholar 

  9. Prento, P. Histochem. J. 27, 906–913 (1995).

    Article  CAS  Google Scholar 

  10. Hayat, M.A. Micron and Microscopica Acta 17, 115–135 (1986).

    Article  CAS  Google Scholar 

  11. Berk, V. et al. Proc. Natl. Acad. Sci. USA 103, 15830–15834 (2006).

    Article  CAS  Google Scholar 

  12. Heras, B. & Martin, J.L. Acta Crystallogr. D Biol. Crystallogr. 61, 1173–1180 (2005).

    Article  Google Scholar 

  13. Lusty, C.L. J. Appl. Cryst. 32, 106–112 (1999).

    Article  CAS  Google Scholar 

  14. Unser, M., Trus, B.L. & Steven, A.C. Ultramicroscopy 23, 39–51 (1987).

    Article  CAS  Google Scholar 

  15. Sander, B. et al. Mol. Cell 24, 267–278 (2006).

    Article  CAS  Google Scholar 

Download references


We thank H. Kohansal, T. Conrad and W. Jahn for expert technical assistance, U. Göringer and M. Brecht for preparing the RNA editing complex, M. Rodnina and K. Gromadski for preparing the 70S ribosome complex, and C. Will for preparing the 17S U2 snRNP. The work was supported by grants from the Federal Ministry of Education and Research, Germany and from the Sixth Framework Programme of the European Union via 3DRepertoire (to H.S.). Work in the laboratory of J.M.P. is supported by Boehringer Ingelheim and by Spots of Excellence of the city of Vienna. N.F. is supported by a Boehringer-Ingelheim fellowship. E.W. is supported by a 'Studienstiftung des deutschen Volkes' fellowship.

Author information

Authors and Affiliations


Corresponding authors

Correspondence to Berthold Kastner or Holger Stark.

Ethics declarations

Competing interests

Max Planck Innovation has files a patent on this method.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3, Supplementary Data, Supplementary Methods (PDF 1220 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Kastner, B., Fischer, N., Golas, M. et al. GraFix: sample preparation for single-particle electron cryomicroscopy. Nat Methods 5, 53–55 (2008).

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