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Three-Dimensional Scanning Transmission Electron Microscopy of Biological Specimens

Published online by Cambridge University Press:  18 January 2010

Niels de Jonge ,
Rachid Sougrat ,
Brian M. Northan  and
Stephen J. Pennycook
Niels de Jonge*
Affiliation:
Vanderbilt University Medical Center, Department of Molecular Physiology and Biophysics, Light Hall 702, Nashville, TN 37232-0615, USA Oak Ridge National Laboratory, Materials Science and Technology Division, 1 Bethel Valley Rd., Oak Ridge, TN 37831-6064, USA
Rachid Sougrat
Affiliation:
Cell Biology and Metabolism Branch, NICHD, National Institute of Health, 18 Library Drive, Bethesda, MD 20892-5430, USA
Brian M. Northan
Affiliation:
Media Cybernetics Inc., 4340 East-West Hwy, Suite 400, Bethesda, MD 20814-4411, USA
Stephen J. Pennycook
Affiliation:
Oak Ridge National Laboratory, Materials Science and Technology Division, 1 Bethel Valley Rd., Oak Ridge, TN 37831-6064, USA
*
Corresponding author. E-mail: niels.de.jonge@vanderbilt.edu
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Abstract

A three-dimensional (3D) reconstruction of the cytoskeleton and a clathrin-coated pit in mammalian cells has been achieved from a focal-series of images recorded in an aberration-corrected scanning transmission electron microscope (STEM). The specimen was a metallic replica of the biological structure comprising Pt nanoparticles 2–3 nm in diameter, with a high stability under electron beam radiation. The 3D dataset was processed by an automated deconvolution procedure. The lateral resolution was 1.1 nm, set by pixel size. Particles differing by only 10 nm in vertical position were identified as separate objects with greater than 20% dip in contrast between them. We refer to this value as the axial resolution of the deconvolution or reconstruction, the ability to recognize two objects, which were unresolved in the original dataset. The resolution of the reconstruction is comparable to that achieved by tilt-series transmission electron microscopy. However, the focal-series method does not require mechanical tilting and is therefore much faster. 3D STEM images were also recorded of the Golgi ribbon in conventional thin sections containing 3T3 cells with a comparable axial resolution in the deconvolved dataset.

Keywords

three-dimensional electron microscopyaberration-corrected STEMbiological electron microscopythin sectionscytoskeletonclathrin-coated pitdeconvolutionnanoparticles
Type
Biological Imaging: Techniques Development and Applications
Information
Microscopy and Microanalysis , Volume 16 , Issue 1 , February 2010 , pp. 54 - 63
Copyright
Copyright © Microscopy Society of America 2010

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