3-D EM exploration of the hepatic microarchitecture – lessons learned from large-volume in situ serial sectioning

To-date serial block-face scanning electron microscopy (SBF-SEM) dominates as the premier technique for generating three-dimensional (3-D) data of resin-embedded biological samples at an unprecedented depth volume. Given the infancy of the technique, limited literature is currently available regardi...

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Vydáno v:Scientific reports Ročník 6; číslo 1; s. 36744
Hlavní autoři: Shami, Gerald John, Cheng, Delfine, Huynh, Minh, Vreuls, Celien, Wisse, Eddie, Braet, Filip
Médium: Journal Article
Jazyk:angličtina
Vydáno: London Nature Publishing Group UK 11.11.2016
Nature Publishing Group
Témata:
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631/1647/245/2221
631/1647/328/1649
631/1647/328/1978
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64
Animals
Bile ducts
Biological samples
Data processing
Electron microscopy
Female
Fixatives
Heavy metals
Humanities and Social Sciences
Imaging, Three-Dimensional
Liver - ultrastructure
Microscopy, Electron, Scanning
multidisciplinary
Parenchymal Tissue - ultrastructure
Rats, Wistar
Sample preparation
Scanning electron microscopy
Science
Sectioning
Signal-To-Noise Ratio
ISSN:2045-2322, 2045-2322
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Shrnutí:To-date serial block-face scanning electron microscopy (SBF-SEM) dominates as the premier technique for generating three-dimensional (3-D) data of resin-embedded biological samples at an unprecedented depth volume. Given the infancy of the technique, limited literature is currently available regarding the applicability of SBF-SEM for the ultrastructural investigation of tissues. Herein, we provide a comprehensive and rigorous appraisal of five different SBF-SEM sample preparation protocols for the large-volume exploration of the hepatic microarchitecture at an unparalleled X, Y and Z resolution. In so doing, we qualitatively and quantitatively validate the use of a comprehensive SBF-SEM sample preparation protocol, based on the application of heavy metal fixatives, stains and mordanting agents. Employing the best-tested SBF-SEM approach, enabled us to assess large-volume morphometric data on murine parenchymal cells, sinusoids and bile canaliculi. Finally, we integrated the validated SBF-SEM protocol with a correlative light and electron microscopy (CLEM) approach. The combination of confocal scanning laser microscopy and SBF-SEM provided a novel way to picture subcellular detail. We appreciate that this multidimensional approach will aid the subsequent research of liver tissue under relevant experimental and disease conditions.
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ISSN:2045-2322
2045-2322
DOI:10.1038/srep36744