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3D d STORM Imaging of Fixed Brain Tissue

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Synapse Development

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1538))

Abstract

Central nervous system tissue contains a high density of synapses each composed of an intricate molecular machinery mediating precise transmission of information. Deciphering the molecular nanostructure of pre- and postsynaptic specializations within such a complex tissue architecture poses a particular challenge for light microscopy. Here, we describe two approaches suitable to examine the molecular nanostructure of synapses at 20–30 nm lateral and 50–70 nm axial resolution within an area of 500 μm × 500 μm and a depth of 0.6 μm to several micrometers. We employ single-molecule localization microscopy (SMLM) on immunolabeled fixed brain tissue slices. tomoSTORM utilizes array tomography to achieve SMLM in 40 nm thick resin-embedded sections. dSTORM of cryo-sectioned slices uses optical sectioning in 0.1–4 μm thick hydrated sections. Both approaches deliver 3D nanolocalization of two or more labeled proteins within a defined tissue volume. We review sample preparation, data acquisition, analysis, and interpretation.

Correspondence may be addressed to either author.

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Author information

Authors and Affiliations

  1. Department of Functional Neuroanatomy, Institute for Anatomy and Cell Biology, Heidelberg University, Im Neuenheimer Feld 307, 69120, Heidelberg, Germany

    Frank Herrmannsdörfer, Benjamin Flottmann, Siddarth Nanguneri, Varun Venkataramani, Heinz Horstmann, Thomas Kuner & Mike Heilemann

  2. Single Molecule Biophysics, Institute of Physical and Theoretical Chemistry, Goethe-University Frankfurt, Max-von-Laue-Str. 7, 60438, Frankfurt, Germany

    Benjamin Flottmann & Mike Heilemann

Authors
  1. Frank Herrmannsdörfer
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  2. Benjamin Flottmann
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  3. Siddarth Nanguneri
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  4. Varun Venkataramani
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  5. Heinz Horstmann
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  6. Thomas Kuner
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  7. Mike Heilemann
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Corresponding author

Correspondence to Thomas Kuner .

Editor information

Editors and Affiliations

  1. Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts, USA

    Alexandros Poulopoulos

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Herrmannsdörfer, F. et al. (2017). 3D d STORM Imaging of Fixed Brain Tissue. In: Poulopoulos, A. (eds) Synapse Development. Methods in Molecular Biology, vol 1538. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6688-2_13

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  • DOI: https://doi.org/10.1007/978-1-4939-6688-2_13

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6686-8

  • Online ISBN: 978-1-4939-6688-2

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