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    High pressure freezing and freeze substitution of Schizosaccharomyces pombe and Saccharomyces cerevisiae for TEM.

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    Authors
    Murray, Stephen M
    Affiliation
    TEM Service Facility, Paterson Institute for Cancer Research, University of Manchester, Manchester, United Kingdom.
    Issue Date
    2008
    
    Metadata
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    Abstract
    The use of standard room temperature chemical fixation protocols for the ultrastructural preservation of yeast and subsequent observation under the electron microscope is fraught with difficulties. Many protocols require the use of enzymatic digestion of the cell wall in order to facilitate the entry of fixatives into the cell interior. Others rely on the use of permanganate-based fixative solutions, which whilst enabling overall preservation of the cell, does require multiple centrifugation, washing, and resuspension steps. This often results in the significant loss of sample volume whilst the use of permanganate can cause extraction of cytoplasmic components. The use of low temperature techniques and in particular high pressure freezing (HPF) and freeze substitution (FS) overcomes many of these problems. With the recent advances in cryotechnologies and in particular the development of commercially available equipment such as the high pressure freezer, the level of ultrastructural preservation attainable in electron microscopy has increased markedly. It is now possible to capture dynamic time sensitive events and to place them in their ultrastructural context with a level of resolution which at the present time can only be achieved with electron microscopy.
    Citation
    High pressure freezing and freeze substitution of Schizosaccharomyces pombe and Saccharomyces cerevisiae for TEM. 2008, 88:3-17 Methods Cell Biol.
    Journal
    Methods in Cell Biology
    URI
    http://hdl.handle.net/10541/55804
    DOI
    10.1016/S0091-679X(08)00401-9
    PubMed ID
    18617025
    Type
    Book chapter
    Language
    en
    ISSN
    0091-679X
    ae974a485f413a2113503eed53cd6c53
    10.1016/S0091-679X(08)00401-9
    Scopus Count
    Collections
    All Paterson Institute for Cancer Research

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