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Live-Cell Quantification of Mitochondrial Functional Parameters

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Visualization Techniques

Part of the book series: Neuromethods ((NM,volume 70))

Abstract

Mitochondria are semi-autonomous organelles, which are central to cellular energy production and signal transduction. Given the tight integration between mitochondrial and cellular physiology, experimental strategies are required to study mitochondrial (dys)function in living cells. For this purpose one can use various chemical and protein-based fluorescent reporter molecules (probes), which are introduced into the cell using specific incubation protocols or transfection techniques. These probes include reporters to monitor mitochondrial membrane potential (Δψ), cytosolic and mitochondrial free calcium concentration (Ca2+), reactive oxygen species (ROS), cytosolic and mitochondrial pH, glucose and ATP. However, proper interpretation and quantification of the above readouts is not trivial. Here, we present our protocol for automated temporal analysis of mitochondrial position in living cells and explain how it can be used for computer-assisted quantification of mitochondrial morphology and Δψ. We further discuss how this approach can be applied for simultaneous quantification of multiple mitochondrial and cellular parameters.

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Acknowledgments

This work was supported by an equipment grant of NWO (Netherlands Organization for Scientific Research, No: 911-02-008), the Dutch Ministry of Economic Affairs (Innovative Onderzoeks Projecten (IOP) Grant: #IGE05003), and by the CSBR (Centres for Systems Biology Research) initiative from NWO (No: CSBR09/013V).

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Authors and Affiliations

  1. Departments of Biochemistry and Pediatrics, Nijmegen Centre for Molecular Life Sciences, Nijmegen Centre of Mitochondrial Disorders, Centre for Systems Biology and Bioenergetics, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

    Marco Nooteboom

  2. Department of Biochemistry, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

    Marleen Forkink

  3. Department of Biochemistry, Nijmegen Centre for Molecular Life Sciences and Centre for Systems Biology and Bioenergetics, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

    Peter H. G. M. Willems & Werner J. H. Koopman

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  1. Marco Nooteboom
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  2. Marleen Forkink
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  3. Peter H. G. M. Willems
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  4. Werner J. H. Koopman
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Corresponding author

Correspondence to Werner J. H. Koopman .

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Editors and Affiliations

  1. School of Medical Sciences, Pharmaceutical Sciences, RMIT University, Plenty Road, Bundoora, 3183, Victoria, Australia

    Emilio Badoer

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Nooteboom, M., Forkink, M., Willems, P.H.G.M., Koopman, W.J.H. (2012). Live-Cell Quantification of Mitochondrial Functional Parameters. In: Badoer, E. (eds) Visualization Techniques. Neuromethods, vol 70. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-897-9_6

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  • DOI: https://doi.org/10.1007/978-1-61779-897-9_6

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-896-2

  • Online ISBN: 978-1-61779-897-9

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