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Preparation of Bacterial Samples for 2-D PAGE

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The Protein Protocols Handbook

Part of the book series: Springer Protocols Handbooks ((SPH))

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Abstract

Sample preparation is a very crucial step in two-dimensional (2-D) gel electro-phoresis in which the proteins of the sample are brought into a state where they can be separated by isoelectric focusing in the first dimension. The proteins must be denatured, reduced, and solubilized, and they must be kept so during electrophoresis without changing their pI. The buffer for this purpose is traditionally called the lysis buffer.

Most bacterial studies aim to solubilize as many proteins as possible to obtain the best possible representation of the total protein content or protein expression under the investigated biological circumstances. However, prefractionation, or the successive application of different chemical reagents, can be used to investigate proteins with certain characteristics. Note that the gels will reflect the proteome of the bacteria at the time the proteins are solubilized and that preceding centrifugations or other manipulations may stress the bacteria, thereby influencing the protein profile.

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Author information

Authors and Affiliations

  1. Novo Nordisk A/S, Maaloev, Denmark

    Brian B. Vandahl

  2. Department of Medical Microbiology and Immunology, University of Aarhus, Aarhus, Denmark

    Gunna Christiansen & Svend Birkelund

Authors
  1. Brian B. Vandahl
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  2. Gunna Christiansen
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  3. Svend Birkelund
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Editor information

Editors and Affiliations

  1. School of Life Sciences, University of Hertfordshire, Hatfield, Hertfordshire, UK

    John M. Walker

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© 2009 Springer Science+Business Media, LLC

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Vandahl, B.B., Christiansen, G., Birkelund, S. (2009). Preparation of Bacterial Samples for 2-D PAGE. In: Walker, J.M. (eds) The Protein Protocols Handbook. Springer Protocols Handbooks. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-59745-198-7_14

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  • DOI: https://doi.org/10.1007/978-1-59745-198-7_14

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60327-474-6

  • Online ISBN: 978-1-59745-198-7

  • eBook Packages: Springer Protocols

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