Targeted nano analysis of water and ions using cryocorrelative light and scanning transmission electron microscopy

Cryo fluorescence imaging coupled with the cryo-EM technique (cryo-CLEM) avoids chemical fixation and embedding in plastic, and is the gold standard for correlated imaging in a close to native state. This multi-modal approach has not previously included elementary nano analysis or evaluation of wate...

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Bibliographic Details
Published in:Journal of structural biology 2012-11, Vol.180 (2), p.352-361
Main Authors: Nolin, Frédérique, Ploton, Dominique, Wortham, Laurence, Tchelidze, Pavel, Balossier, Gérard, Banchet, Vincent, Bobichon, Hélène, Lalun, Nathalie, Terryn, Christine, Michel, Jean
Format: Article
Language:English
Subjects:
Correlative microscopy
Cryoelectron Microscopy - methods
Cryomicroscopy
EDXS
Ions - chemistry
Microscopy, Electron, Scanning Transmission - methods
Nano analysis
STEM
Water - chemistry
Water content
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Summary:Cryo fluorescence imaging coupled with the cryo-EM technique (cryo-CLEM) avoids chemical fixation and embedding in plastic, and is the gold standard for correlated imaging in a close to native state. This multi-modal approach has not previously included elementary nano analysis or evaluation of water content. We developed a new approach allowing analysis of targeted in situ intracellular ions and water measurements at the nanoscale (EDXS and STEM dark field imaging) within domains identified by examination of specific GFP-tagged proteins. This method allows both water and ions- fundamental to cell biology- to be located and quantified at the subcellular level. We illustrate the potential of this approach by investigating changes in water and ion content in nuclear domains identified by GFP-tagged proteins in cells stressed by Actinomycin D treatment and controls. The resolution of our approach was sufficient to distinguish clumps of condensed chromatin from surrounding nucleoplasm by fluorescence imaging and to perform nano analysis in this targeted compartment.
ISSN:1047-8477
1095-8657
DOI:10.1016/j.jsb.2012.08.011