Immunoelectron Microscopy of Cryofixed Freeze-Substituted Saccharomyces cerevisiae

  • Jindriska Fiserova
  • Martin W. Goldberg
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 657)

Abstract

Immunolabelling electron microscopy is a challenging technique with demands for perfect ultrastructural and antigen preservation. High-pressure freezing offers an ideal way to fix cellular structure. However, its use for immunolabelling has remained limited because of the low frequency of labelling due to loss of protein antigenicity or accessibility. Here we present a protocol for immunogold labelling of the yeast Saccharomyces cerevisiae that gives specific and multiple labelling while keeping the finest structural details. We use the protocol to reveal the organisation of individual nuclear pore complex proteins and the position of transport factors in the yeast S. cerevisiae in relation to actual transport events.

Key words

Yeast nuclear transport transmission electron microscopy freeze substitution lowicryl immunostaining 

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Jindriska Fiserova
    • 1
  • Martin W. Goldberg
    • 1
  1. 1.School of Biological and Biomedical SciencesDurham UniversityDurhamUK

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