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Genetically Encoded Reporters to Monitor Hypoxia

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Hypoxia

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2755))

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Abstract

Hypoxia resulting from an imbalance of oxygen availability and consumption defines a metabolic cellular state with a profound impact on developmental processes, tissue maintenance, and the development of pathologies. Fluorescence imaging using genetically encoded reporters enables hypoxia and oxygen imaging with cellular resolution. Thereby unrestricted visualization of hypoxic cells and regions essentially relies on the availability of oxygen-independent fluorescent proteins like UnaG, isolated from the Japanese freshwater eel. Here, we describe the application of recently developed members of a UnaG-based hypoxia reporter family to visualize oxygenation patterns by in vitro live-cell imaging and during the ex vivo analysis of intracranial xenografted tumors. Thus, the generation of stably transfected transgenic tumor cell lines, the in vitro calibration of the genetically encoded sensors, the surgical procedures for orthotopic xenografting of tumors in mice, and workflows for the respective sample preparation and microscopy are outlined.

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Acknowledgments

This work was supported by Raghu Erapaneedi (EIMI, WWU) by contributing a microscopy image (Fig. 6b), by Dirk Reinhardt (EIMI, WWU) by performing the intracranial tumor injections, and by Martin Stehling (Flow cytometry-Unit, MPI Münster) with expert technical support. We gratefully acknowledge funding by the Deutsche Forschungsgemeinschaft (SFB1450/1-431460824) and by the Studienstiftung des Deutschen Volkes to Nadine Bauer.

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Authors and Affiliations

  1. European Institute for Molecular Imaging, University of Münster, Münster, Germany

    Nadine Bauer & Friedemann Kiefer

  2. Max Planck Institute for Molecular Biomedicine, Münster, Germany

    Nadine Bauer & Friedemann Kiefer

Authors
  1. Nadine Bauer
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  2. Friedemann Kiefer
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Corresponding author

Correspondence to Friedemann Kiefer .

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Editors and Affiliations

  1. Department of Oncology Johns Hopkins, University School of Medicine, Baltimore, MD, USA

    Daniele M. Gilkes

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© 2024 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Bauer, N., Kiefer, F. (2024). Genetically Encoded Reporters to Monitor Hypoxia. In: Gilkes, D.M. (eds) Hypoxia. Methods in Molecular Biology, vol 2755. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3633-6_1

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  • DOI: https://doi.org/10.1007/978-1-0716-3633-6_1

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3632-9

  • Online ISBN: 978-1-0716-3633-6

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