Microtubules in Plant Cells: Strategies and Methods for Immunofluorescence, Transmission Electron Microscopy, and Live Cell Imaging

  • Katherine Celler
  • Miki Fujita
  • Eiko Kawamura
  • Chris Ambrose
  • Klaus Herburger
  • Andreas Holzinger
  • Geoffrey O. Wasteneys
Part of the Methods in Molecular Biology book series (MIMB, volume 1365)


Microtubules (MTs) are required throughout plant development for a wide variety of processes, and different strategies have evolved to visualize and analyze them. This chapter provides specific methods that can be used to analyze microtubule organization and dynamic properties in plant systems and summarizes the advantages and limitations for each technique. We outline basic methods for preparing samples for immunofluorescence labeling, including an enzyme-based permeabilization method, and a freeze-shattering method, which generates microfractures in the cell wall to provide antibodies access to cells in cuticle-laden aerial organs such as leaves. We discuss current options for live cell imaging of MTs with fluorescently tagged proteins (FPs), and provide chemical fixation, high-pressure freezing/freeze substitution, and post-fixation staining protocols for preserving MTs for transmission electron microscopy and tomography.

Key words

Microtubules EB1 GFP Kinesin MAP4 MBD MOR1 ARK1 Electron tomography Live cell imaging Correlative light and electron microscopy Immunofluorescence 



This chapter has been supported by Austrian Science Fund (FWF) project P24242-B16 to AH and funding from the Natural Sciences and Engineering Research Council and the Canadian Institutes of Health Research to GOW. The Thieme Verlag KG is acknowledged for their kind permission to reproduce Fig. 1b, c. Figure 6a–c is reproduced with permission of the National Academy of Sciences of the United States of America, and Dr. Jennifer Lippincott-Schwartz, National Institute of Child Health and Human Development, Bethesda, MD. Elsevier Inc. for reproduction of Fig. 10.


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Katherine Celler
    • 1
  • Miki Fujita
    • 1
  • Eiko Kawamura
    • 2
  • Chris Ambrose
    • 3
  • Klaus Herburger
    • 4
  • Andreas Holzinger
    • 4
  • Geoffrey O. Wasteneys
    • 1
  1. 1.Department of BotanyThe University of British ColumbiaVancouverCanada
  2. 2.Western College of Veterinary MedicineUniversity of SaskatchewanSaskatoonCanada
  3. 3.Department of BiologyUniversity of SaskatchewanSaskatoonCanada
  4. 4.Functional Plant Biology, Institute of BotanyUniversity of InnsbruckInnsbruckAustria

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