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Three-dimensional distributions of elements in biological samples by energy-filtered electron tomography.

Abstract : By combining electron tomography with energy-filtered electron microscopy, we have shown the feasibility of determining the three-dimensional distributions of phosphorus in biological specimens. Thin sections of the nematode, Caenorhabditis elegans were prepared by high-pressure freezing, freeze-substitution and plastic embedding. Images were recorded at energy losses above and below the phosphorus L2,3 edge using a post-column imaging filter operating at a beam energy of 120 keV. The unstained specimens exhibited minimal contrast in bright-field images. After it was determined that the specimen was sufficiently thin to allow two-window ratio imaging of phosphorus, pairs of pre-edge and post-edge images were acquired in series over a tilt range of +/-55 degrees at 5 degrees increments for two orthogonal tilt axes. The projected phosphorus distributions were aligned using the pre-edge images that contained inelastic contrast from colloidal gold particles deposited on the specimen surface. A reconstruction and surface rendering of the phosphorus distribution clearly revealed features 15-20 nm in diameter, which were identified as ribosomes distributed along the stacked membranes of endoplasmic reticulum and in the cytoplasm. The sensitivity of the technique was estimated at < 35 phosphorus atoms per voxel based on the known total ribosomal phosphorus content of approximately 7000 atoms. Although a high electron dose of approximately 10(7)e/nm2 was required to record two-axis tilt series, specimens were sufficiently stable to allow image alignment and tomographic reconstruction.
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https://www.hal.inserm.fr/inserm-00078802
Contributor : Dominique Laurent-Maquin <>
Submitted on : Wednesday, June 7, 2006 - 3:26:57 PM
Last modification on : Thursday, February 7, 2019 - 5:13:40 PM

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Richard Leapman, Elisabeth Kocsis, Gu Zhang, Thomas Talbot, Patrice Laquerriere. Three-dimensional distributions of elements in biological samples by energy-filtered electron tomography.. Ultramicroscopy, Elsevier, 2004, 100, pp.115-25. ⟨10.1016/j.ultramic.2004.03.002⟩. ⟨inserm-00078802⟩

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