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Preparation of Mammalian Tissue Samples for Two-Dimensional Electrophoresis

  • Frank A. Witzmann
Protocol
Part of the Springer Protocols Handbooks book series (SPH)

Abstract

The true power of two-dimensional gel electrophoresis (2-DE) requires the careful preparation of protein samples to minimize sample variability and maximize solubilization. This is of particular relevance when 2-DE is relied upon to characterize the differential expression of mammalian tissue proteomes in large experiments with large numbers of samples. One of the weaknesses of the 2-DE approach relates to its depth of field-that is, its limited ability to resolve reproducibly those proteins with extreme isoelectric points (pI) (e.g., >3 and <9), hydrophobicity, and low abundance. Second, the initial step in 2-DE of protein mixtures, isoelectric focusing, is susceptible to a number of problems that cause variability in the final protein pattern, interferences that must be avoided. Hence, a simple yet reproducible method of preparing cell and tissue samples for consistent 2-DE results, involving as little manipulation as possible, is of utmost importance.

Keywords

Carrier Ampholyte Isocyanic Acid Solubilized Sample Fisher Sonic Primary Hepatocyte Cell Culture 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Humana Press Inc., Totowa, NJ 2005

Authors and Affiliations

  • Frank A. Witzmann
    • 1
  1. 1.Department of Cellular & Integrative PhysiologyIndiana University School of MedicineIndianapolis

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