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  Ethane-freezing/methanol-fixation of cell monolayers: a procedure for improved preservation of structure and antigenicity for light and electron microscopies

Neuhaus, E. M., Horstmann, H., Almers, W., Maniak, M., & Soldati, T. (1998). Ethane-freezing/methanol-fixation of cell monolayers: a procedure for improved preservation of structure and antigenicity for light and electron microscopies. Journal of Structural Biology, 121(3), 326-342. doi:10.1006/jsbi.1998.3971.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002B-7282-4 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002B-7283-2
Genre: Journal Article
Alternative Title : Ethane-freezing/methanol-fixation of cell monolayers: a procedure for improved preservation of structure and antigenicity for light and electron microscopies

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JStructBiol_121_1998_326.pdf (Any fulltext), 4MB
 
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 Creators:
Neuhaus, Eva Maria1, Author              
Horstmann, Heinz1, Author              
Almers, Wolfhard1, Author              
Maniak, Markus, Author
Soldati, Thierry2, Author              
Affiliations:
1Department of Molecular Cell Research, Max Planck Institute for Medical Research, Max Planck Society, ou_1497703              
2Department of Biomedical Optics, Max Planck Institute for Medical Research, Max Planck Society, ou_1497699              

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 Abstract: In order to dissect at the ultrastructural level the morphology of highly dynamic processes such as cell motility, membrane trafficking events, and organelle movements, it is necessary to fix/stop time-dependent events in the millisecond range. Ideally, immunoelectron microscopical labeling experiments require the availability of high-affinity antibodies and accessibility to all compartments of the cell. The biggest challenge is to define an optimum between significant preservation of the antigenicity in the fixed material without compromising the intactness of fine structures. Here, we present a procedure which offers an opportunity to unify preparation of cell monolayers for immunocytochemistry in fluorescence and electron microscopy. This novel strategy combines a rapid ethane-freezing technique with a low temperature methanol-fixation treatment (EFMF) and completely avoids chemical fixatives. It preserves the position and delicate shape of cells and organelles and leads to improved accessibility of the intracellular antigens and to high antigenicity preservation. We illustrate the establishment of this procedure usingDictyostelium discoideum,a powerful model organism to study molecular mechanisms of membrane trafficking and cytoskeleton.

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Language(s): eng - English
 Dates: 1998
 Publication Status: Published in print
 Pages: 17
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 Table of Contents: -
 Rev. Type: Peer
 Degree: -

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Title: Journal of Structural Biology
  Abbreviation : J. Struct. Biol.
Source Genre: Journal
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Publ. Info: Orlando, Fla. : Academic Press
Pages: - Volume / Issue: 121 (3) Sequence Number: - Start / End Page: 326 - 342 Identifier: ISSN: 1047-8477
CoNE: https://pure.mpg.de/cone/journals/resource/954922650160