Rigid Embedding of Fixed and Stained, Whole, Millimeter-Scale Specimens for Section-free 3D Histology by Micro-Computed Tomography

Alex Y. Lin, Yifu Ding, Daniel J. Vanselow, Spencer R. Katz, Maksim A. Yakovlev, Darin P. Clark, David Mandrell, Jean E. Copper, Damian B. Van Rossum, Keith Cheng

Research output: Contribution to journalArticle

Abstract

For over a hundred years, the histological study of tissues has been the gold standard for medical diagnosis because histology allows all cell types in every tissue to be identified and characterized. Our laboratory is actively working to make technological advances in X-ray micro-computed tomography (micro-CT) that will bring the diagnostic power of histology to the study of full tissue volumes at cellular resolution (i.e., an X-ray Histo-tomography modality). Toward this end, we have made targeted improvements to the sample preparation pipeline. One key optimization, and the focus of the present work, is a straightforward method for rigid embedding of fixed and stained millimeter-scale samples. Many of the published methods for sample immobilization and correlative micro-CT imaging rely on placing the samples in paraffin wax, agarose, or liquids such as alcohol. Our approach extends this work with custom procedures and the design of a 3-dimensional printable apparatus to embed the samples in an acrylic resin directly into polyimide tubing, which is relatively transparent to X-rays. Herein, sample preparation procedures are described for the samples from 0.5 to 10 mm in diameter, which would be suitable for whole zebrafish larvae and juveniles, or other animals and tissue samples of similar dimensions. As proof of concept, we have embedded the specimens from Danio, Drosophila, Daphnia, and a mouse embryo; representative images from 3-dimensional scans for three of these samples are shown. Importantly, our methodology leads to multiple benefits including rigid immobilization, long-term preservation of laboriously-created resources, and the ability to re-interrogate samples.

Original languageEnglish (US)
JournalJournal of visualized experiments : JoVE
Issue number140
DOIs
StatePublished - Oct 17 2018

Fingerprint

Histology
Tomography
Tissue
X rays
Immobilization
X Ray Tomography
Daphnia
Paraffin waxes
Acrylic Resins
X-Ray Microtomography
Waxes
Zebrafish
Tubing
Polyimides
Paraffin
Sepharose
Drosophila
Acrylics
Larva
Animals

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Lin, Alex Y. ; Ding, Yifu ; Vanselow, Daniel J. ; Katz, Spencer R. ; Yakovlev, Maksim A. ; Clark, Darin P. ; Mandrell, David ; Copper, Jean E. ; Van Rossum, Damian B. ; Cheng, Keith. / Rigid Embedding of Fixed and Stained, Whole, Millimeter-Scale Specimens for Section-free 3D Histology by Micro-Computed Tomography. In: Journal of visualized experiments : JoVE. 2018 ; No. 140.
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Rigid Embedding of Fixed and Stained, Whole, Millimeter-Scale Specimens for Section-free 3D Histology by Micro-Computed Tomography. / Lin, Alex Y.; Ding, Yifu; Vanselow, Daniel J.; Katz, Spencer R.; Yakovlev, Maksim A.; Clark, Darin P.; Mandrell, David; Copper, Jean E.; Van Rossum, Damian B.; Cheng, Keith.

In: Journal of visualized experiments : JoVE, No. 140, 17.10.2018.

Research output: Contribution to journalArticle

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AU - Ding, Yifu

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AU - Katz, Spencer R.

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AU - Clark, Darin P.

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AU - Cheng, Keith

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