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Extended Methods for 2D Confinement

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Cell Migration in Three Dimensions

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2608))

Abstract

Physical confinement in microfluidic devices has become a common technique to induce and study cell migration in a large range of cell types. Confined migration was previously understudied due to the limitations of 2D migration assays but has emerged as an important mode of migration in the past decade. Furthermore, confinement improves the quality of the imaging and simplifies the analysis of trajectories by confining migration to the plane of acquisition. Protocols described in this chapter relate to methods extending the previously published 2D confinement technique. First, we explain a method to increase the complexity of the confinement chamber by microfabricating nanometer-sized PDMS grooves on the bottom surface, usually used for contact guidance studies. Then, we describe a method to perform the confinement on cells embedded inside a μm-thin 3D collagen gel. Finally, we describe an alternative method to confine cells based on agarose, so that cells can be fixed or drug perfused while being confined, which is currently not possible in the 2D confinement silicone-based device.

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Acknowledgments

We thank Sergii Rudiuk (Ecole Normale Superieure, Paris) for helping with the AFM profiling of the CD-R PDMS replica. We thank Jin Wei (Ecole Normale Superieure, Paris) for helping with the SEM imaging of the CD-R PDMS replica. This work was supported by a French Agence Nationale de la Recherche (ANR) grant to MP (ANR-19-CE13-0030). This work has also received the support of Institut Pierre-Gilles de Gennes-IPGG (équipement d’excellence, “investissements d’avenir,” program ANR-10-EQPX-34) and laboratoire d’excellence, “investissements d’avenir” program ANR-10-IDEX-0001-02 PSL and ANR-10-LABX-31. We also thank the staff members at the Nikon Imaging Center at Institut Curie and the technological platform at Institut Pierre-Gilles de Gennes (IPGG) for providing equipment and technical assistance. JMGA has received funding from Inserm (ITMO Cancer FDV2016), Fondation ARC pour la recherche sur le cancer (DOC20190508743), and a short-term EMBO fellowship (7873).

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Authors and Affiliations

  1. Institut Pierre Gilles de Gennes, PSL Research University, Paris, France

    Juan M. García-Arcos, Kevin Gateau, Larisa Venkova & Matthieu Piel

  2. Current affiliation, Department of Biochemistry and Swiss National Centre of Competence in Research, Chemical Biology, University of Geneva, Geneva, Switzerland

    Juan M. García-Arcos

  3. Ecole Normale Supérieure Paris-Saclay, Gif-sur-Yvette, France

    Kevin Gateau

  4. Current affiliation, Institute of Biochemistry and Cellular Genetics, UMR 5095, CNRS, University of Bordeaux, Bordeaux, France

    Larisa Venkova

Authors
  1. Juan M. García-Arcos
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  2. Kevin Gateau
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  3. Larisa Venkova
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  4. Matthieu Piel
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Corresponding author

Correspondence to Matthieu Piel .

Editor information

Editors and Affiliations

  1. Department of Medical Oncology, Cancer Center Amsterdam, Amsterdam University Medical Center, Amsterdam, The Netherlands

    Coert Margadant

1 Electronic Supplementary Material(s)

Video 1

Representative time-lapse images from confocal fluorescence spinning disk microscopy of HeLa Kyoto cells Lifeact-mCherry MYH9-GFP confined under 3 μm over a nanogrooved surface treated with 0,5 mg/ml PLL-g-PEG (ZIP 95994 kb)

Video 2

Representative z-stack images from confocal fluorescence spinning disk microscopy of HeLa Kyoto cells Lifeact-mCherry MYH9-GFP confined under 10 and under 3 μm embedded in a collagen type I gel of 1 mg/mL (ZIP 4981 kb)

Agarose_holder_fluorodish.stl

3D print model for the agarose confiner to be used with glass-bottom FluoroDish™ Petri dishes (WPI) (ZIP 33 kb)

Agarose_holder_TTPdish.stl

3D print model for the agarose confiner to be used with TPP™ Polystyrene Petri dishes (ZIP 31 kb)

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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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García-Arcos, J.M., Gateau, K., Venkova, L., Piel, M. (2023). Extended Methods for 2D Confinement. In: Margadant, C. (eds) Cell Migration in Three Dimensions. Methods in Molecular Biology, vol 2608. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2887-4_5

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  • DOI: https://doi.org/10.1007/978-1-0716-2887-4_5

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2886-7

  • Online ISBN: 978-1-0716-2887-4

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