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Visualization and Quantitative Analysis of the Actin Cytoskeleton Upon B Cell Activation

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B Cell Receptor Signaling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1707))

Abstract

The formation of the immunological synapse upon B cell activation critically depends on the rearrangement of the submembranous actin cytoskeleton. Polymerization of actin monomers into filaments provides the force required for B cell spreading on the antigen-presenting cell (APC). Interestingly, the actin network also participates in cellular signaling at multiple levels. Fluorescence microscopy plays a critical role in furthering our understanding of the various functions of the cytoskeleton, and has become an important tool in the studies on B cell activation. The actin cytoskeleton can be tracked in live cells with various fluorescent probes binding to actin, or in fixed cells typically with phalloidin staining. Here, we present the usage of TIRF microscopy and an image analysis workflow for studying the overall density and organization of the actin network upon B cell spreading on antigen-coated glass, a widely used model system for the formation of the immunological synapse.

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Acknowledgments

Imaging was performed at the Cell Imaging Core, Turku Centre for Biotechnology, University of Turku and Åbo Akademi University. We thank Markku Saari and Jouko Sandholm from the Cell Imaging Core for their help with the microscopes, and the whole Mattila lab for their critical comments to the manuscript. Turku University Hospital (TYKS) is acknowledged for the support in basic laboratory materials. This work was supported by the Academy of Finland (grant ID: 25700, 296684 and 307313 for PKM, and 286712 for VŠ), University of Turku Graduate School (UTUGS) (for MV), VSSHP research support (for PKM), as well as Sigrid Juselius (for PKM), Jane and Aatos Erkko (for PKM), Magnus Ehrnrooth (for PKM and VŠ) and Finnish Cultural (for VŠ) foundations.

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Correspondence to Pieta K. Mattila .

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Šuštar, V., Vainio, M., Mattila, P.K. (2018). Visualization and Quantitative Analysis of the Actin Cytoskeleton Upon B Cell Activation. In: Liu, C. (eds) B Cell Receptor Signaling. Methods in Molecular Biology, vol 1707. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7474-0_18

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  • DOI: https://doi.org/10.1007/978-1-4939-7474-0_18

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7473-3

  • Online ISBN: 978-1-4939-7474-0

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