Correlative light- and electron microscopy with chemical tags

Correlative microscopy incorporates the specificity of fluorescent protein labeling into high-resolution electron micrographs. Several approaches exist for correlative microscopy, most of which have used the green fluore
Correlative microscopy incorporates the specificity of fluorescent protein labeling into high-resolution electron micrographs. Several approaches exist for correlative microscopy, most of which have used the green fluorescent protein (GFP) as the label for light microscopy. Here we use chemical tagging and synthetic fluorophores instead, in order to achieve protein-specific labeling, and to perform multicolor imaging. We show that synthetic fluorophores preserve their post-embedding fluorescence in the presence of uranyl acetate. Post-embedding fluorescence is of such quality that the specimen can be prepared with identical protocols for scanning electron microscopy (SEM) and transmission electron microscopy (TEM); this is particularly valuable when singular or otherwise difficult samples are examined. We show that synthetic fluorophores give bright, well-resolved signals in super-resolution light microscopy, enabling us to superimpose light microscopic images with a precision of up to 25 nm in the x-y plane on electron micrographs. To exemplify the preservation quality of our new method we visualize the molecular arrangement of cadherins in adherens junctions of mouse epithelial cells.
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Author:Mario Perkovic, Michael Kunz, Ulrike Endesfelder, Stefanie Bunse, Christoph Wigge, Zhou Yu, Victor-Valentin Hodirnau, Margot P. Scheffer, Anja Seybert, Sebastian Malkusch, Erin Schuman, Mike Heilemann, Achilleas S. Frangakis
URN:urn:nbn:de:hebis:30:3-348115
DOI:http://dx.doi.org/10.1016/j.jsb.2014.03.018
ISSN:1095-8657
ISSN:1047-8477
Pubmed Id:http://www.ncbi.nlm.nih.gov/pubmed?term=24698954
Parent Title (English):Journal of structural biology
Publisher:Elsevier
Place of publication:San Diego, Calif.
Document Type:Article
Language:English
Year of Completion:2014
Date of first Publication:2014/03/31
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2014/10/27
Tag:Adherens junctions; Correlative electron and light microscopy; Electron tomography
Volume:186
Issue:2
Pagenumber:9
First Page:205
Last Page:213
Note:
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
HeBIS PPN:366621548
Institutes:Biochemie und Chemie
MPI für Hirnforschung
Dewey Decimal Classification:540 Chemie und zugeordnete Wissenschaften
Sammlungen:Universitätspublikationen
Licence (German):License LogoCreative Commons - Namensnennung-Nicht kommerziell-Keine Bearbeitung 3.0

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