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Cell Imaging Techniques pp 295–312Cite as

Mica Functionalization for Imaging of DNA and Protein-DNA Complexes with Atomic Force Microscopy

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 931))

Abstract

Surface preparation is a key step for reliable and reproducible imaging of DNA and protein-DNA complexes with atomic force microscopy (AFM). This article describes the approaches for chemical functionalization of the mica surface. One approach utilizes 3-aminopropyl-trietoxy silane (APTES), enabling one to obtain a smooth surface termed AP-mica. This surface binds nucleic acids and nucleoprotein complexes in a wide range of ionic strengths, in the absence of divalent cations and in a broad range of pH. Another method utilizes aminopropyl silatrane (APS) to yield an APS-mica surface. The advantage of APS-mica compared with AP-mica is the ability to obtain reliable and reproducible time-lapse images in aqueous solutions. The chapter describes the methodologies for the preparation of AP-mica and APS-mica surfaces and the preparation of samples for AFM imaging. The protocol for synthesis and purification of APS is also provided. The applications are illustrated with a number of examples.

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Acknowledgements

The authors thank the members of the Lyubchenko lab for their contribution to different parts of the paper and T. Zaikova for useful comments on the APS synthesis. The work is supported by grants to YLL from the DOE (DE-FG02-08ER64579), NATO (SfP 983204), NIH (P01 GM091743, 1R01 GM096039), the Nebraska Research Initiative (NRI), National Institutes of Health Grants (1P01GM091743-01A1 and 1 R01 GM096039-01A1), US Department of Energy Grant DE-FG02-08ER64579, National Science Foundation (EPS—1004094), and the Nebraska Research Initiative grant to Y.L.L.

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE, USA

    Luda S. Shlyakhtenko & Yuri L. Lyubchenko

  2. Cepheid, Bothell, WA, USA

    Alexander A. Gall

Authors
  1. Luda S. Shlyakhtenko
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  2. Alexander A. Gall
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  3. Yuri L. Lyubchenko
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Corresponding author

Correspondence to Yuri L. Lyubchenko .

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Editors and Affiliations

  1. College of Medicine, Professor, Department of Pathology, University of Vermont, Beaumont Ave 89, Burlington, 05405-1742, Vermont, USA

    Douglas J. Taatjes

  2. , Abt. Zell- und Molekularpathologie, Universität Zürich, Schmelzbergstr. 12, Zürich, 8091, Switzerland

    Jürgen Roth

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Shlyakhtenko, L.S., Gall, A.A., Lyubchenko, Y.L. (2012). Mica Functionalization for Imaging of DNA and Protein-DNA Complexes with Atomic Force Microscopy. In: Taatjes, D., Roth, J. (eds) Cell Imaging Techniques. Methods in Molecular Biology, vol 931. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-056-4_14

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  • DOI: https://doi.org/10.1007/978-1-62703-056-4_14

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  • Publisher Name: Humana Press, Totowa, NJ

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