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Nanostructure analysis using spatially modulated illumination microscopy

Nature Protocols volume 2, pages 2640–2646 (2007)Cite this article

Abstract

We describe the usage of the spatially modulated illumination (SMI) microscope to estimate the sizes (and/or positions) of fluorescently labeled cellular nanostructures, including a brief introduction to the instrument and its handling. The principle setup of the SMI microscope will be introduced to explain the measures necessary for a successful nanostructure analysis, before the steps for sample preparation, data acquisition and evaluation are given. The protocol starts with cells already attached to the cover glass. The protocol and duration outlined here are typical for fixed specimens; however, considerably faster data acquisition and in vivo measurements are possible.

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Figure 1: SMI setup.
Figure 2: SMI measurements of fluorescence in situ hybridization-labeled centromere 8 regions in VH7 cells.
Figure 3: SMI measurements of the Ki-67 protein14 (a proliferation marker protein) using a standard antibody protocol in VH7 cells.

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Acknowledgements

This work was sponsored by grants from the German Research Foundation (DFG CR 60/23-1+2) and by the European Commission (EU LSHG-CT-2003-503259 and MEIF-CT-2006-041827).

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Author notes

  1. Institute for Pharmacy and Molecular Biotechnology, Universität Heidelberg, D-69120 Heidelberg, Germany

Authors and Affiliations

  1. Kirchhoff Institut für Physik, Universität Heidelberg, INF 227, Heidelberg, D-69120, Germany

    David Baddeley, Claudia Batram, Yanina Weiland, Christoph Cremer & Udo J Birk

  2. Institute of Molecular Biophysics, The Jackson Laboratory, Bar Harbor, 04609, Maine, USA

    Christoph Cremer

  3. Randall Division of Cell & Molecular Biophysics, King's College London, London, SE1 1UL, UK

    Udo J Birk

  4. Institute of Electronic Structure & Laser—FORTH, PO Box 1527, Vassilika Vouton, 71110, Heraklion, Greece

    Udo J Birk

Authors
  1. David Baddeley
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  2. Claudia Batram
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  3. Yanina Weiland
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  4. Christoph Cremer
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  5. Udo J Birk
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Corresponding author

Correspondence to David Baddeley.

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Supplementary Manual

Basic equipment setup and alignment procedure (PDF 94 kb)

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Baddeley, D., Batram, C., Weiland, Y. et al. Nanostructure analysis using spatially modulated illumination microscopy. Nat Protoc 2, 2640–2646 (2007). https://doi.org/10.1038/nprot.2007.399

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