Skip to main content

Automated Image Acquisition for Low-Dose STEM at Atomic Resolution

  • Andreas Mittelberger (a1), Christian Kramberger (a1), Christoph Hofer (a1), Clemens Mangler (a1) and Jannik C. Meyer (a1)...

Beam damage is a major limitation in electron microscopy that becomes increasingly severe at higher resolution. One possible route to circumvent radiation damage, which forms the basis for single-particle electron microscopy and related techniques, is to distribute the dose over many identical copies of an object. For the acquisition of low-dose data, ideally no dose should be applied to the region of interest before the acquisition of data. We present an automated approach that can collect large amounts of data efficiently by acquiring images in a user-defined area-of-interest with atomic resolution. We demonstrate that the stage mechanics of the Nion UltraSTEM, combined with an intelligent algorithm to move the sample, allow the automated acquisition of atomically resolved images from micron-sized areas of a graphene substrate. Moving the sample stage automatically in a regular pattern over the area-of-interest enables the collection of data from pristine sample regions without exposing them to the electron beam before recording an image. Therefore, it is possible to obtain data with minimal dose (no prior exposure during focusing), which is only limited by the minimum signal needed for data processing. This enables us to minimize beam-induced damage in the sample and to acquire large data sets within a reasonable amount of time.

Corresponding author
* Corresponding author.
Hide All
Banhart, F., Kotakoski, J. & Krasheninnikov, A.V. (2011). Structural defects in graphene. ACS Nano 5(1), 2641.
Bartesaghi, A., Merk, A., Banerjee, S., Matthies, D., Wu, X., Milne, J.L.S. & Subramaniam, S. (2015). 2.2 Å resolution cryo-EM structure of β-galactosidase in complex with a cell-permeant inhibitor. Science 348(6239), 11471151.
Cheng, Y. (2015). Single-particle Cryo-EM at crystallographic resolution. Cell 161(3), 450457.
Cheng, Y., Grigorieff, N., Penczek, P. & Walz, T. (2015). A primer to single-particle cryo-electron microscopy. Cell 161(3), 438449.
Dierksen, K., Typke, D., Hegerl, R., Koster, A. & Baumeister, W. (1992). Towards automatic electron tomography. Ultramicroscopy 40(1), 7187.
Egerton, R.F. (2012). Mechanisms of radiation damage in beam-sensitive specimens, for TEM accelerating voltages between 10 and 300 kV. Microsc Res Tech 75(11), 15501556.
Frank, J., Goldfarb, W., Eisenberg, D. & Baker, T.S. (1978). Reconstruction of glutamine synthetase using computer averaging. Ultramicroscopy 3(C), 283290.
Fultz, B. & Howe, J.M. (2008). Transmission electron microscopy and diffractometry of materials . Berlin, Heidelberg, New York: Springer-Verlag.
Koster, A., Chen, H., Sedat, J. & Agard, D. (1992). Automated microscopy for electron tomography. Ultramicroscopy 46(1–4), 207227.
Kotakoski, J., Krasheninnikov, A.V., Kaiser, U. & Meyer, J.C. (2011). From point defects in graphene to two-dimensional amorphous carbon. Phys Rev Lett 106(10), 105505.
Kotakoski, J., Mangler, C. & Meyer, J.C. (2014). Imaging atomic-level random walk of a point defect in graphene. Nat Commun 5, 3991.
Kramberger, C. & Meyer, J.C. (2016). Progress in structure recovery from low dose exposures: Mixed molecular adsorption, exploitation of symmetry and reconstruction from the minimum signal level. Ultramicroscopy 170, 6068.
Mastronarde, D.N. (2005). Automated electron microscope tomography using robust prediction of specimen movements. J Struct Biol 152(1), 3651.
Meyer, J.C., Eder, F., Kurasch, S., Skakalova, V., Kotakoski, J., Park, H.J., Roth, S., Chuvilin, A., Eyhusen, S., Benner, G., Krasheninnikov, A.V. & Kaiser, U. (2012). Accurate measurement of electron beam induced displacement cross sections for single-layer graphene. Phys Rev Lett 108(19), 196102.
Meyer, J.C., Kisielowski, C., Erni, R., Rossell, M.D., Crommie, M.F. & Zettl, A. (2008). Direct Imaging of Lattice Atoms and Topological Defects in Graphene Membranes. Nano Letters 8(11), 35823586.
Meyer, J.C., Kotakoski, J. & Mangler, C. (2014). Atomic structure from large-area, low-dose exposures of materials: A new route to circumvent radiation damage. Ultramicroscopy 145, 1321.
Robertson, A.W., Allen, C.S., Wu, Y.a., He, K., Olivier, J., Neethling, J., Kirkland, A.I. & Warner, J.H. (2012). Spatial control of defect creation in graphene at the nanoscale. Nat Commun 3, 1144.
Shi, J., Williams, D.R. & Stewart, P.L. (2008). A Script-Assisted Microscopy (SAM) package to improve data acquisition rates on FEI Tecnai electron microscopes equipped with Gatan CCD cameras. J Struct Biol 164(1), 166169.
Suloway, C., Pulokas, J., Fellmann, D., Cheng, A., Guerra, F., Quispe, J., Stagg, S., Potter, C.S. & Carragher, B. (2005). Automated molecular microscopy: The new Leginon system. J Struct Biol 151(1), 4160.
Zhang, P., Beatty, A., Milne, J.L. & Subramaniam, S. (2001). Automated data collection with a Tecnai 12 electron microscope: Applications for molecular imaging by cryomicroscopy. J Struct Biol 135(3), 251261.
Zhang, P., Borgnia, M.J., Mooney, P., Shi, D., Pan, M., O’Herron, P., Mao, A., Brogan, D., Milne, J.L. & Subramaniam, S. (2003). Automated image acquisition and processing using a new generation of 4Kx4K CCD cameras for cryo electron microscopic studies of macromolecular assemblies. J Struct Biol 143(2), 135144.
Zhang, X., Jin, L., Fang, Q., Hui, W.H. & Zhou, Z.H. (2010). 3.3 Å cryo-EM structure of a nonenveloped virus reveals a priming mechanism for cell entry. Cell 141(3), 472482.
Recommend this journal

Email your librarian or administrator to recommend adding this journal to your organisation's collection.

Microscopy and Microanalysis
  • ISSN: 1431-9276
  • EISSN: 1435-8115
  • URL: /core/journals/microscopy-and-microanalysis
Please enter your name
Please enter a valid email address
Who would you like to send this to? *



Altmetric attention score

Full text views

Total number of HTML views: 15
Total number of PDF views: 121 *
Loading metrics...

Abstract views

Total abstract views: 318 *
Loading metrics...

* Views captured on Cambridge Core between 23rd May 2017 - 22nd April 2018. This data will be updated every 24 hours.