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Cited by 6Advanced Electron Microscopy Techniques and ApplicationsAdvanced Materials Characterization TechniquesElectron and X-Ray Spectroscopy Techniques

Contrast-Enhanced Tissue Processing of Fibrillin-Rich Elastic Fibres for 3D Visualization by Volume Scanning Electron Microscopy

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Lewis, Philip N., Young, Robert D., Souza, R. B., Quantock, Andrew J., Meek, Keith M.·2021·Methods and Protocols
DOI 10.3390/mps4030056PMC PMC8395850
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Abstract

Elastic fibres constitute an important component of the extracellular matrix and currently are the subject of intensive study in order to elucidate their assembly, function and involvement in cell–matrix interactions and disease. However, few studies to date have investigated the 3D architecture of the elastic fibre system in bulk tissue. We describe a protocol for preparation of tissue samples, including primary fixation and backscatter electron contrast-enhancement steps, through dehydration into stable resin-embedded blocks for volume electron microscopy. The use of low molecular weight tannic acid and alcoholic lead staining are critical stages in this procedure. Block preparation by ultramicrotomy and evaporative metal coating prior to microscopical examination are also described. We present images acquired from serial block face scanning electron microscopy of cornea and aorta showing target structures clearly differentiated from cells and other matrix components. The processing method imparts high contrast to fibrillin-containing elastic fibres, thus facilitating their segmentation and rendering into 3D reconstructions by image analysis software from large serial image datasets.

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  • At the centre of the cornea, elastic fibres exist as thin bundles of fibrillin-rich microfibrils.

    “whereas at the centre of the cornea, they exist as thin bundles of fibrillin-rich microfibrils.”

    From Structural control of corneal transparency, refractive power and dynamics (2025) · cites this paper as [30]

    The Introduction (page unavailable) says microfibrils occur in the cornea, and Expected Results (page unavailable) describes elastic fibres in the human corneal stroma. Neither passage establishes that fibres specifically at the centre of the cornea form thin bundles of fibrillin-rich microfibrils. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • Elastic fibres in the limbus contain fibrillin microfibrils surrounding an elastin core.

    “in the limbus they contain fibrillin microfibrils surrounding an elastin core”

    From Structural control of corneal transparency, refractive power and dynamics (2025) · cites this paper as [30]

    The Introduction (page unavailable) describes elastic fibres generally as fibrillin-rich microfibrils surrounding an elastin core. Expected Results (page unavailable) identifies fibrillin-containing elastic sheets in the human corneal limbus, but does not establish that those limbal structures have an elastin core. (LLM, conf=0.83)

    Experimental — accuracy not yet measured

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Research works printed but not linked: 25

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Sorted by citations

Authors
Lewis, P. N., Young, R. D., Souza, R. B., Quantock, A. J., & Meek, K. M.
Affiliations
  • Cardiff University (GB)
  • Universidade de São Paulo (BR)
Corresponding author
Lewis, Philip N.
ORCID
Venue
Methods and Protocols
Publisher
MDPI AG
Published
2021-08-15
Volume
4
Issue
3
Pages
56
ISSN
2409-9279 (ISSN-L)
DOI
10.3390/mps4030056
PMC
PMC8395850
OpenAlex
W3193754908
Cited by
6
References
0
Citations by year
2025: 1 · 2024: 2 · 2023: 3
FWCI
0.52
Type
journal article
Language
en
Open access
Licence
Locations
  • Methods and Protocols (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • ORCA Online Research @Cardiff (Cardiff University) (acceptedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
Medical Research Council (MR/SO37829/1)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Advanced Electron Microscopy Techniques and Applications
  • Physical Sciences › Engineering › Biomedical Engineering › Advanced Materials Characterization Techniques
  • Physical Sciences › Materials Science › Surfaces, Coatings and Films › Electron and X-Ray Spectroscopy Techniques
Keywords
elastic fibers, fibrillin, extracellular matrix, tissue processing, tannic acid, resin embedding, serial block-face scanning electron microscopy, 3D reconstruction, cornea, aorta, ultramicrotomy

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