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© Thierry Blisnick & Philippe Bastin, Institut Pasteur
Bloodstream Trypanosoma brucei cell
Publication : Micron (Oxford, England : 1993)

Scanning transmission electron microscopy through-focal tilt-series on biological specimens

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Micron (Oxford, England : 1993) - 29 May 2015

Trepout S, Messaoudi C, Perrot S, Bastin P, Marco S

Link to Pubmed [PMID] – 26093182

Micron 2015 May;77:9-15

Since scanning transmission electron microscopy can produce high signal-to-noise ratio bright-field images of thick (≥500nm) specimens, this tool is emerging as the method of choice to study thick biological samples via tomographic approaches. However, in a convergent-beam configuration, the depth of field is limited because only a thin portion of the specimen (from a few nanometres to tens of nanometres depending on the convergence angle) can be imaged in focus. A method known as through-focal imaging enables recovery of the full depth of information by combining images acquired at different levels of focus. In this work, we compare tomographic reconstruction with the through-focal tilt-series approach (a multifocal series of images per tilt angle) with reconstruction with the classic tilt-series acquisition scheme (one single-focus image per tilt angle). We visualised the base of the flagellum in the protist Trypanosoma brucei via an acquisition and image-processing method tailored to obtain quantitative and qualitative descriptors of reconstruction volumes. Reconstructions using through-focal imaging contained more contrast and more details for thick (≥500nm) biological samples.