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Cytometric Analysis of DNA Damage: Phosphorylation of Histone H2AX as a Marker of DNA Double-Strand Breaks (DSBs)

  • Toshiki Tanaka
  • Dorota Halicka
  • Frank Traganos
  • Zbigniew Darzynkiewicz
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 523)

Abstract

Phosphorylation of histone H2AX on Ser 139 is a sensitive reporter of DNA damage, particularly if the damage involves induction of DNA double-strand breaks (DSBs). Phosphorylated H2AX has been named γH2AX and its presence in the nucleus can be detected immunocytochemically. Multiparameter analysis of γH2AX immunofluorescence by flow or laser-scanning cytometry allows one to measure extent of DNA damage in individual cells and to correlate it with their position in the cell cycle and induction of apoptosis. This chapter presents the protocols and outlines applications of multiparameter cytometry in analysis of H2AX phosphorylation as a reporter of the presence of DSBs.

Key words

γH2AX H2AX phosphorylation DNA double-strand breaks Multiparameter flow cytometry Laser-scanning cytometry Immunocytochemistry Apoptosis 

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Toshiki Tanaka
    • 1
    • 2
  • Dorota Halicka
    • 2
  • Frank Traganos
    • 2
  • Zbigniew Darzynkiewicz
    • 2
  1. 1.Department of Surgery and Clinical Science, Division of Chest SurgeryGraduate School of Medicine, Yamaguchi UniversityYamaguchiJapan
  2. 2.Department of PathologyBrander Cancer Research Institute, New York Medical CollegeValhallaUSA

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