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In Vivo Light Sheet Fluorescence Microscopy of Calcium Oscillations in Arabidopsis thaliana

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Book cover Calcium Signalling

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

Abstract

Calcium imaging in plants requires a high-resolution microscope, able to perform volumetric acquisition in a few seconds, inducing as low photobleaching and phototoxicity as possible to the sample. Light sheet fluorescence microscopy offers these capabilities, with the further chance to mount the sample in vertical position, mimicking the plant’s growth and physiological conditions.

A protocol for plant preparation and mounting in a light sheet microscope is presented. First, the growth of Arabidopsis thaliana in a sample holder compatible with light sheet microscopy is described. Then, the requirements for sample alignment and image acquisition are detailed. Finally, the image processing steps to analyze calcium oscillations are discussed, with particular emphasis on ratiometric calcium imaging in Arabidopsis root hairs.

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Acknowledgments

This work was supported by Laserlab-Europe [EU-H2020 654148] and by Università degli Studi di Milano [PIANO DI SVILUPPO DI ATENEO 2016] to A.Co. NRA was a beneficiary of a fellowship from the European Commission within the framework of the “SUSTAIN-T Project of the Erasmus Mundus Programme, Action 2—STRAND 1, Lot 7, Latin America.”

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Authors and Affiliations

  1. Dipartimento di Fisica, Politecnico di Milano, Milan, Italy

    Neli Romano Armada, Alessia Candeo, Gianluca Valentini & Andrea Bassi

  2. INIQUI y Facultad de Ingeniería, Universidad Nacional de Salta, Salta, Argentina

    Neli Romano Armada

  3. Dipartimento di Bioscienze, Università degli studi di Milano, Milan, Italy

    Fabrizio Gandolfo Doccula & Alex Costa

  4. Istituto di Fotonica e Nanotecnologie, Consiglio Nazionale delle Ricerche, Milan, Italy

    Gianluca Valentini & Andrea Bassi

Authors
  1. Neli Romano Armada
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  2. Fabrizio Gandolfo Doccula
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  3. Alessia Candeo
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  4. Gianluca Valentini
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  5. Alex Costa
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  6. Andrea Bassi
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Corresponding author

Correspondence to Alex Costa .

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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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Romano Armada, N., Doccula, F.G., Candeo, A., Valentini, G., Costa, A., Bassi, A. (2019). In Vivo Light Sheet Fluorescence Microscopy of Calcium Oscillations in Arabidopsis thaliana . 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_8

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  • DOI: https://doi.org/10.1007/978-1-4939-9018-4_8

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9017-7

  • Online ISBN: 978-1-4939-9018-4

  • eBook Packages: Springer Protocols

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