Clem methods for studying primary cilia

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

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

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.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages193-202
Number of pages10
Volume1454
DOIs
StatePublished - 2016

Publication series

NameMethods in Molecular Biology
Volume1454
ISSN (Print)10643745

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Keywords

  • Axoneme
  • CLEM
  • MEF
  • Primary cilia
  • SEM
  • Superresolution

ASJC Scopus subject areas

  • Medicine(all)
  • Molecular Biology
  • Genetics

Cite this

Macaluso, F. P., Perumal, G., Kolstrup, J., & Satir, P. (2016). Clem methods for studying primary cilia. In Methods in Molecular Biology (Vol. 1454, pp. 193-202). (Methods in Molecular Biology; Vol. 1454). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-3789-9_12