3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy

Matthew R.G. Russell, Thomas R. Lerner, Jemima J. Burden, David O. Nkwe, Annegret Pelchen-Matthews, Marie Charlotte Domart, Joanne Durgan, Anne Weston, Martin L. Jones, Christopher J. Peddie, Raffaella Carzaniga, Oliver Florey, Mark Marsh, Maximiliano G. Gutierrez, Lucy M. Collinson

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here, we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy, and 3D image processing and analysis. We demonstrate successful application of the workflow to three studies, each aiming to better understand complex and dynamic biological processes, including bacterial and viral infections of cultured cells and formation of entotic cell-in-cell structures commonly observed in tumours. Our workflow revealed new insight into the replicative niche of Mycobacterium tuberculosis in primary human lymphatic endothelial cells, HIV-1 in human monocytederived macrophages, and the composition of the entotic vacuole. The broad application of this 3D CLEM technique will make it a useful addition to the correlative imaging toolbox for biomedical research.

Original languageEnglish
Pages (from-to)278-291
Number of pages14
JournalJournal of Cell Science
Volume130
Issue number1
DOIs
Publication statusPublished - Jan 1 2017

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Workflow
Electron Scanning Microscopy
Cultured Cells
Electron Microscopy
Light
Cell-in-Cell Formation
Biological Phenomena
Virus Diseases
Vacuoles
Mycobacterium tuberculosis
Fluorescence Microscopy
Bacterial Infections
HIV-1
Biomedical Research
Endothelial Cells
Cell Culture Techniques
Macrophages
Neoplasms

All Science Journal Classification (ASJC) codes

  • Cell Biology

Cite this

Russell, M. R. G., Lerner, T. R., Burden, J. J., Nkwe, D. O., Pelchen-Matthews, A., Domart, M. C., ... Collinson, L. M. (2017). 3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy. Journal of Cell Science, 130(1), 278-291. https://doi.org/10.1242/jcs.188433
Russell, Matthew R.G. ; Lerner, Thomas R. ; Burden, Jemima J. ; Nkwe, David O. ; Pelchen-Matthews, Annegret ; Domart, Marie Charlotte ; Durgan, Joanne ; Weston, Anne ; Jones, Martin L. ; Peddie, Christopher J. ; Carzaniga, Raffaella ; Florey, Oliver ; Marsh, Mark ; Gutierrez, Maximiliano G. ; Collinson, Lucy M. / 3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy. In: Journal of Cell Science. 2017 ; Vol. 130, No. 1. pp. 278-291.
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Russell, MRG, Lerner, TR, Burden, JJ, Nkwe, DO, Pelchen-Matthews, A, Domart, MC, Durgan, J, Weston, A, Jones, ML, Peddie, CJ, Carzaniga, R, Florey, O, Marsh, M, Gutierrez, MG & Collinson, LM 2017, '3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy', Journal of Cell Science, vol. 130, no. 1, pp. 278-291. https://doi.org/10.1242/jcs.188433

3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy. / Russell, Matthew R.G.; Lerner, Thomas R.; Burden, Jemima J.; Nkwe, David O.; Pelchen-Matthews, Annegret; Domart, Marie Charlotte; Durgan, Joanne; Weston, Anne; Jones, Martin L.; Peddie, Christopher J.; Carzaniga, Raffaella; Florey, Oliver; Marsh, Mark; Gutierrez, Maximiliano G.; Collinson, Lucy M.

In: Journal of Cell Science, Vol. 130, No. 1, 01.01.2017, p. 278-291.

Research output: Contribution to journalArticle

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AU - Russell, Matthew R.G.

AU - Lerner, Thomas R.

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AU - Pelchen-Matthews, Annegret

AU - Domart, Marie Charlotte

AU - Durgan, Joanne

AU - Weston, Anne

AU - Jones, Martin L.

AU - Peddie, Christopher J.

AU - Carzaniga, Raffaella

AU - Florey, Oliver

AU - Marsh, Mark

AU - Gutierrez, Maximiliano G.

AU - Collinson, Lucy M.

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