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From Tube to Structure: SPA Cryo-EM Workflow Using Apoferritin as an Example

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Structural Proteomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2305))

Abstract

In this chapter, we present an overview of a standard protocol to achieve structure determination at high resolution by Single Particle Analysis cryogenic Electron Microscopy using apoferritin as a standard sample. The purified apoferritin is applied to a glow-discharged support and then flash frozen in liquid ethane. The prepared grids are loaded into the electron microscope and checked for particle spreading and ice thickness. The microscope alignments are performed and the data collection session is setup for an overnight data collection. The collected movies containing two-dimensional images of the apoferritin sample are then processed to obtain a high-resolution three-dimensional reconstruction.

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Authors and Affiliations

  1. The Netherlands Centre for Electron Nanoscopy (NeCEN), Leiden University, Leiden, The Netherlands

    Christoph A. Diebolder, Rebecca S. Dillard & Ludovic Renault

Authors
  1. Christoph A. Diebolder
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  2. Rebecca S. Dillard
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  3. Ludovic Renault
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Corresponding author

Correspondence to Ludovic Renault .

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Editors and Affiliations

  1. The Rosalind Franklin Institute, Harwell Science Campus, Didcot, UK

    Raymond J. Owens

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Diebolder, C.A., Dillard, R.S., Renault, L. (2021). From Tube to Structure: SPA Cryo-EM Workflow Using Apoferritin as an Example. In: Owens, R.J. (eds) Structural Proteomics. Methods in Molecular Biology, vol 2305. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1406-8_12

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  • DOI: https://doi.org/10.1007/978-1-0716-1406-8_12

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1405-1

  • Online ISBN: 978-1-0716-1406-8

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