Multiparameter Cell Cycle Analysis

  • James W. Jacobberger
  • R. Michael Sramkoski
  • Tammy Stefan
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 699)

Abstract

Cell cycle-related cytometry and analysis is an essential experimental paradigm for the cell biology of yeast, mammalian, and drosophila cells. Methods have not changed much for many years. The most common is DNA content analysis, which has been well-published and reviewed. Next most common is analysis of 5-bromo-2-deoxyuridine (BrdU) incorporation, detected by specific antibodies – also well-published and reviewed. A new measurement approach to S phase labeling utilizes 5′-ethynyl-2′-deoxyuridine (EdU) incorporation and a chemical reaction to label substituted DNA. The approach is new, but published work indicates that it is equivalent to BrdU incorporation. Finally, multiple antibody labeling to detect epitopes on cell cycle-regulated proteins is the most complex of the cytometric cell cycle assays, requiring knowledge of the chemistry of fixation, the biochemistry of antibody–antigen reactions, and spectral compensation. Because all of this knowledge is relatively well presented, methodologically, in many papers and reviews, this chapter presents a bare-bones Methods section for one mammalian cell type and an extended Notes section, focusing on aspects that are problematic or not well described in the literature.

Key words

Cell division cycle Mitosis Antibodies Intracellular antigens Cyclins Cell states 

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • James W. Jacobberger
    • 1
  • R. Michael Sramkoski
    • 1
  • Tammy Stefan
    • 1
  1. 1.Cytometry and Imaging Microscopy Core, Case Comprehensive Cancer CenterCase Western Reserve UniversityClevelandUSA

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