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Quantitative Immunofluorescence Analysis of Nucleolus-Associated Chromatin

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Book cover The Nucleolus

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1455))

Abstract

The nuclear distribution of eu- and heterochromatin is nonrandom, heterogeneous, and dynamic, which is mirrored by specific spatiotemporal arrangements of histone posttranslational modifications (PTMs). Here we describe a semiautomated method for the analysis of histone PTM localization patterns within the mammalian nucleus using confocal laser scanning microscope images of fixed, immunofluorescence stained cells as data source. The ImageJ-based process includes the segmentation of the nucleus, furthermore measurements of total fluorescence intensities, the heterogeneity of the staining, and the frequency of the brightest pixels in the region of interest (ROI). In the presented image analysis pipeline, the perinucleolar chromatin is selected as primary ROI, and the nuclear periphery as secondary ROI.

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Acknowledgements

This work was supported by the DFG SFB960 program.

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Authors and Affiliations

  1. Biochemistry Center Regensburg, University of Regensburg, Universitaetsstr. 31, 93053, Regensburg, Germany

    Stefan Dillinger & Attila Németh

Authors
  1. Stefan Dillinger
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  2. Attila Németh
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Corresponding authors

Correspondence to Stefan Dillinger or Attila Németh .

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Editors and Affiliations

  1. Department of Biochemistry III, University of Regensburg, Regensburg, Germany

    Attila Németh

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Dillinger, S., Németh, A. (2016). Quantitative Immunofluorescence Analysis of Nucleolus-Associated Chromatin. In: Németh, A. (eds) The Nucleolus. Methods in Molecular Biology, vol 1455. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3792-9_5

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  • DOI: https://doi.org/10.1007/978-1-4939-3792-9_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3790-5

  • Online ISBN: 978-1-4939-3792-9

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