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High-throughput imaging of heterogeneous cell organelles with an X-ray laser

Nature Photonics volume 8, pages 943–949 (2014)Cite this article

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Abstract

We overcome two of the most daunting challenges in single-particle diffractive imaging: collecting many high-quality diffraction patterns on a small amount of sample and separating components from mixed samples. We demonstrate this on carboxysomes, which are polyhedral cell organelles that vary in size and facilitate up to 40% of Earth's carbon fixation. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min with the Linac Coherent Light Source running at 120 Hz. We separate different structures directly from the diffraction data and show that the size distribution is preserved during sample delivery. We automate phase retrieval and avoid reconstruction artefacts caused by missing modes. We attain the highest-resolution reconstructions on the smallest single biological objects imaged with an X-ray laser to date. These advances lay the foundations for accurate, high-throughput structure determination by flash-diffractive imaging and offer a means to study structure and structural heterogeneity in biology and elsewhere.

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Figure 1: The experimental set-up to image aerosolized carboxysomes.
Figure 2: Sample injection.
Figure 3: Computational purification of carboxysomes using low-resolution fit parameters and the autocorrelation function.
Figure 4: Measured size distribution of single carboxysome particles in the gas phase.
Figure 5: Computational purification of carboxysomes on the basis of size and shape.
Figure 6: Reconstructed image of a single carboxysome compared to the image of an icosahedron with uniform density.

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Acknowledgements

This work was supported by the Swedish Research Council, the Knut and Alice Wallenberg Foundation, the European Research Council, the Röntgen-Ångström Cluster and Stiftelsen Olle Engkvist Byggmästare. Portions of this research were carried out at the Linac Coherent Light Source, a national user facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences. The authors thank the scientific and technical staff of the LCLS for support. The authors thank the CAMP collaboration for giving access to their experimental set-up and for supporting the experiment at the LCLS. The authors also acknowledge the Max Planck Society for funding the development and operation of the CAMP instrument.

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Author notes

  1. Max F. Hantke and Dirk Hasse: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Cell and Molecular Biology, Laboratory of Molecular Biophysics, Uppsala University, Husargatan 3 (Box 596), Uppsala, SE-751 24, Sweden

    Max F. Hantke, Dirk Hasse, Filipe R. N. C. Maia, Tomas Ekeberg, Katja John, Martin Svenda, Nicusor Timneanu, Daniel S. D. Larsson, Gijs van der Schot, Gunilla H. Carlsson, Margareta Ingelman, Jakob Andreasson, Daniel Westphal, M. Marvin Seibert, Kerstin Mühlig, Janos Hajdu & Inger Andersson

  2. NERSC, Lawrence Berkeley National Laboratory, Berkeley, 94720, California, USA

    Filipe R. N. C. Maia

  3. Centre for BioImaging Sciences, National University of Singapore, 14 Science Drive 4, 117557, Singapore, Singapore

    N. Duane Loh

  4. ARC Centre of Excellence for Coherent X-ray Science, School of Physics, The University of Melbourne, Victoria, 3010, Australia

    Andrew V. Martin

  5. Center for Free-Electron Laser Science, DESY, Notkestrasse 85, Hamburg, 22607, Germany

    Mengning Liang, Francesco Stellato, Richard A. Kirian, Daniel Rolles, Sascha Epp, Henry N. Chapman & Anton Barty

  6. INFN and Physics Department, University of Rome ‘Tor Vergata’, Via della Ricerca Scientifica 1, Rome, 00133, Italy

    Francesco Stellato

  7. LCLS, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025, California, USA

    Daniel P. DePonte, M. Marvin Seibert, Sebastian Schorb, Ken Ferguson, Christoph Bostedt, Sebastian Carron & John D. Bozek

  8. PNSensor GmbH, Römerstrasse 28, München, 80803, Germany

    Robert Hartmann

  9. Max Planck Institute for Extraterrestrial Physics, Giessenbachstrasse, Garching, 85741, Germany

    Nils Kimmel

  10. Department of Physics, Kansas State University, 331 Cardwell Hall, Kansas, 66506, Manhattan, USA

    Artem Rudenko

  11. European XFEL GmbH, Albert-Einstein-Ring 19, Hamburg, 22761, Germany

    Janos Hajdu

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  1. Max F. Hantke
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Contributions

J.H., I.A., M.F.H., F.R.N.C.M. and T.E. developed the imaging concept and conceived the experiment. M.F.H., F.R.N.C.M., T.E., A.B., N.D.L., A.M., G.V.D.S. and D.L. developed ideas and software to process the diffraction data. D.H., K.J., G.H.C., M.S., M.I. and I.A. prepared and characterized carboxysomes for the study. J.H., B.I., D.P.D., R.A.K., M.S., J.A., M.M.S. and D.W. developed and operated the sample injector. J.D.B., C.B., S.C., N.T., M.S. and M.M.S. operated the beamline at the LCLS. R.H. and N.K. operated the pnCCD detectors. M.F.H., F.R.N.C.M., T.E., N.D.L., G.V.D.S., A.B., J.A., M.M.S., M.S., M.L., F.S., D.R., A.R., S.E., H.N.C. and J.H. characterized the imaging apparatus and carried out the experiment. M.F.H., F.R.N.C.M. and T.E. processed the data. M.F.H., F.R.N.C.M., I.A. and J.H. analysed the results and wrote the manuscript with input from the others.

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Correspondence to Janos Hajdu.

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Hantke, M., Hasse, D., Maia, F. et al. High-throughput imaging of heterogeneous cell organelles with an X-ray laser. Nature Photon 8, 943–949 (2014). https://doi.org/10.1038/nphoton.2014.270

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