Dehydration as alternative sample preparation for soft X-ray tomography.
X-ray absorption
cell anatomy
dehydration
tomography
Journal
Journal of microscopy
ISSN: 1365-2818
Titre abrégé: J Microsc
Pays: England
ID NLM: 0204522
Informations de publication
Date de publication:
09 2023
09 2023
Historique:
revised:
26
06
2023
received:
06
04
2023
accepted:
09
07
2023
medline:
9
8
2023
pubmed:
12
7
2023
entrez:
11
7
2023
Statut:
ppublish
Résumé
Soft X-ray tomography (SXT) is an imaging technique to visualise whole cells without fixation, staining, and sectioning. For SXT imaging, cells are cryopreserved and imaged at cryogenic conditions. Such 'near-to-native' state imaging is in high demand and initiated the development of the laboratory table-top SXT microscope. As many laboratories do not have access to cryogenic equipment, we asked ourselves whether SXT imaging is feasible on dry specimens. This paper shows how the dehydration of cells can be used as an alternative sample preparation to obtain ultrastructure information. We compare different dehydration processes on mouse embryonic fibroblasts in terms of ultrastructural preservation and shrinkage. Based on this analysis, we chose critical point (CPD) dried cells for SXT imaging. In comparison to cryopreserved and air-dried cells, CPD dehydrated cells show high structural integrity although with about 3-7 times higher X-ray absorption for cellular organelles. As the difference in X-ray absorption values between organelles is preserved, 3D anatomy of CPD-dried cells can be segmented and analysed, demonstrating the applicability of CPD-dried sample preparation for SXT imaging. LAY DESCRIPTION: Soft X-ray tomography (SXT) is an imaging technique that allows to see the internal structures of cells without the need for special treatments like fixation or staining. Typically, SXT imaging involves freezing and imaging cells at very low temperatures. However, since many labs lack the necessary equipment, we explored whether SXT imaging could be done on dry samples instead. We compared different dehydration methods and found that critical point drying (CPD) was the most promising for SXT imaging. CPD-dried cells showed high structural integrity, although they absorbed more X-rays than hydrated cells, demonstrating that CPD-dried sample preparation is a viable alternative for SXT imaging.
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
248-255Informations de copyright
© 2023 The Authors. Journal of Microscopy published by John Wiley & Sons Ltd on behalf of Royal Microscopical Society.
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