Immunoelectron Microscopy of Cryofixed Freeze-Substituted Yeast Saccharomyces cerevisiae
Language English Country United States Media print
Document type Journal Article
Grant support
BB/E015735/1
Biotechnology and Biological Sciences Research Council - United Kingdom
BB/G011818/1
Biotechnology and Biological Sciences Research Council - United Kingdom
- Keywords
- Freeze substitution, Immunostaining, Lowicryl, Nuclear, Transmission electron microscopy, Transport, Yeast,
- MeSH
- Staining and Labeling methods MeSH
- Epoxy Resins chemistry MeSH
- Gene Expression MeSH
- Tissue Fixation methods MeSH
- Fixatives chemistry MeSH
- Glutaral chemistry MeSH
- Microscopy, Immunoelectron methods MeSH
- Immunohistochemistry methods MeSH
- Nuclear Pore Complex Proteins genetics metabolism MeSH
- Cryopreservation methods MeSH
- Microtomy MeSH
- Freeze Substitution methods MeSH
- Antibodies chemistry MeSH
- Saccharomyces cerevisiae Proteins genetics metabolism MeSH
- Saccharomyces cerevisiae metabolism ultrastructure MeSH
- Tissue Embedding methods MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Epoxy Resins MeSH
- Fixatives MeSH
- Glutaral MeSH
- Nuclear Pore Complex Proteins MeSH
- Antibodies MeSH
- Saccharomyces cerevisiae Proteins MeSH
Immunolabeling electron microscopy is a challenging technique with demands for perfect ultrastructural and antigen preservation. High-pressure freezing offers an excellent way to fix cellular structure. However, its use for immunolabeling has remained limited because of the low frequency of labeling due to loss of protein antigenicity or accessibility. Here we present a protocol for immunogold labeling of the yeast Saccharomyces cerevisiae that gives specific and multiple labeling while keeping the finest structural details. We use the protocol to reveal the organization of individual nuclear pore complex proteins and the position of transport factors in the yeast Saccharomyces cerevisiae in relation to actual transport events.
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