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Calcium Signalling pp 31–42Cite as

Measuring Ca2+ Levels in Subcellular Compartments with Genetically Encoded GFP-Based Indicators

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1925))

Abstract

Ca2+ homeostasis is crucial for the entire life of eukaryotic cells from the beginning to the end. Mishandling in Ca2+ homeostasis is indeed linked with a large number of pathological conditions. Thus, the possibility to specifically monitor cellular calcium fluxes in different subcellular compartments represents a key tool to deeply understand the mechanisms involved in cellular dysfunctions. To cope with this need, several Ca2+ indicators have been developed allowing to accurately measure both basal Ca2+ concentration and agonist-induced Ca2+ signals in a wide spectrum of organelles. Among these, the genetically encoded GFP-based indicators are routinely used to measure Ca2+ transients thanks to their ability to change their spectral properties in response to Ca2+ binding. In this chapter, we will describe a protocol that utilizes the GCaMP6f probe targeted to mitochondria (4mtGCaMP) to measure mitochondrial calcium levels in resting conditions in HeLa cells. This method allows to easily and quickly register alterations of mitochondrial Ca2+ homeostasis in different cell populations and experimental settings, representing a precious tool to unravel the pathological pathways leading to pathogenic conditions.

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Authors and Affiliations

  1. Department of Biomedical Sciences, University of Padua, Padua, Italy

    Mattia Vicario & Tito Calì

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  1. Mattia Vicario
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  2. Tito Calì
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Correspondence to Tito Calì .

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Editors and Affiliations

  1. Department of Biomedical Sciences, University of Padua, Padua, Italy

    Anna Raffaello

  2. Department of Biomedical Sciences, University of Padua, Padua, Italy

    Denis Vecellio Reane

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Vicario, M., Calì, T. (2019). Measuring Ca2+ Levels in Subcellular Compartments with Genetically Encoded GFP-Based Indicators. In: Raffaello, A., Vecellio Reane, D. (eds) Calcium Signalling. Methods in Molecular Biology, vol 1925. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9018-4_3

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  • DOI: https://doi.org/10.1007/978-1-4939-9018-4_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9017-7

  • Online ISBN: 978-1-4939-9018-4

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