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Unbiased Quantification of Golgi Scattering and Golgi–Centrosome Association

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Golgi

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2557))

Abstract

The vertebrate Golgi complex is a large dynamic organelle which undergoes morphological changes and fragmentation both as a part of normal physiological dynamics and under disease conditions. The Golgi is known to have a functionally important relationship with the centrosome. The extent of the spatial association between these two organelles varies in a dynamic and regulated manner. It is essential to have a reliable unbiased approach to evaluate Golgi volume, Golgi extension/scattering in the 3D cell space, and spatial association of the Golgi with the centrosome. It is also important that each of these features is evaluated by a simple metric, one measurement per cell, so that the variability and deviations in the cell population can be easily assessed. Here, we present an approach to analyze confocal microscopy image stacks to easily measure Golgi volume, scattering, and association with the centrosome. The approach is based on a custom MATLAB script, provided here as a supplement, and also uses widely available software (ImageJ and/or Imaris). The output of the script is a table with the following parameters: Golgi volume in voxels, Golgi volume in μm3, “Golgi–Golgi” distance (averaged distance between all Golgi voxels), Golgi–centrosome distance (averaged distance between each Golgi voxel and the nearest mother centriole), and centrosome–centrosome distance (for cells with duplicated centrosome, the distance between the mother centrioles). The approach can also be applied to analyze distribution of any fluorescently- labeled structure within a cell and its association with the centrosome or any single point within the cell volume.

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Acknowledgments

This work was supported by the National Institutes of Health (NIH) grants R35-GM127098 (to I.K.), R01-DK106228 (to I.K.), and R35-GM130298 (to A.K.). K.B.F. was supported by an NIH training grant R25-GM062459 “Initiative for Maximize Student Diversity” (Sealy, PI). We utilized the Flow Cytometry Shared Resource supported by the Vanderbilt Ingram Cancer Center for cell sorting.

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Authors and Affiliations

  1. Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN, USA

    Keyada B. Frye, Xiaodong Zhu & Irina Kaverina

  2. Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA

    Xiaodong Zhu

  3. Wadsworth Center, New York State Department of Health, Albany, NY, USA

    Alexey Khodjakov

Authors
  1. Keyada B. Frye
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  2. Xiaodong Zhu
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  3. Alexey Khodjakov
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  4. Irina Kaverina
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Corresponding author

Correspondence to Irina Kaverina .

Editor information

Editors and Affiliations

  1. Department of Molecular, Cellular and Developmental Biology, University of Michigan, Ann Arbor, MI, USA

    Yanzhuang Wang

  2. Department of Physiology and Cell Biology, University of Arkansas for Medical Sciences, Little Rock, AR, USA

    Vladimir V Lupashin

  3. Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA

    Todd R. Graham

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Frye, K.B., Zhu, X., Khodjakov, A., Kaverina, I. (2023). Unbiased Quantification of Golgi Scattering and Golgi–Centrosome Association. In: Wang, Y., Lupashin, V.V., Graham, T.R. (eds) Golgi. Methods in Molecular Biology, vol 2557. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2639-9_31

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  • DOI: https://doi.org/10.1007/978-1-0716-2639-9_31

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2638-2

  • Online ISBN: 978-1-0716-2639-9

  • eBook Packages: Springer Protocols

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