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CLEM Methods for Studying Primary Cilia

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Cilia

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1454))

Abstract

CLEM (correlated light and electron microscope) imaging is a highly useful technique for examining primary cilia. With CLEM, it is possible to determine the distribution of tagged proteins along the ciliary membrane and axoneme with high precision. Scanning electron microscopy (SEM) permits measurement of ciliary length and orientation in relation to nearby cellular structures in a 3D image; in optimal cases, this can be combined with superresolution microscopy of selected ciliary components as they enter or leave the cilium. This chapter discusses CLEM methods. In the method described in detail, samples are completely processed for sequential fluorescence and SEM observation. This method is ideal for robust antibody localization and minimizes image manipulation in correlating the fluorescent and SEM images. Alternative methods prepare samples for fluorescence imaging followed by processing for SEM then observation in the SEM. This method is ideal for optimal fluorescence imaging, particularly live cell imaging.

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Authors and Affiliations

  1. Analytical Imaging Facility, Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Forchheimer Building, Room 602, 1300 Morris Park Ave., Bronx, NY, 10461, USA

    Frank P. Macaluso, Geoffrey S. Perumal, Johan Kolstrup & Peter Satir

Authors
  1. Frank P. Macaluso
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  2. Geoffrey S. Perumal
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  3. Johan Kolstrup
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  4. Peter Satir
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Corresponding author

Correspondence to Peter Satir .

Editor information

Editors and Affiliations

  1. Albert Einstein College of Medicine, Yeshiva University, Bronx, New York, USA

    Peter Satir

  2. Department of Biology, University of Copenhagen, Copenhagen, Denmark

    Søren Tvorup Christensen

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© 2016 Springer Science+Business Media New York

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Macaluso, F.P., Perumal, G.S., Kolstrup, J., Satir, P. (2016). CLEM Methods for Studying Primary Cilia. In: Satir, P., Christensen, S. (eds) Cilia. Methods in Molecular Biology, vol 1454. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3789-9_12

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  • DOI: https://doi.org/10.1007/978-1-4939-3789-9_12

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3787-5

  • Online ISBN: 978-1-4939-3789-9

  • eBook Packages: Springer Protocols

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