Cytometric analysis of DNA damage: phosphorylation of histone H2AX as a marker of DNA double-strand breaks (DSBs).
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 gammaH2AX and its presence in the nucleus can be detected immunocytochemically. Multiparameter analysis of gammaH2AX 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.
Tanaka
Toshiki
T
Department of Surgery and Clinical Science, Division of Chest Surgery, Graduate School of Medicine, Yamaguchi University, Yamaguchi, Japan.
Halicka
Dorota
D
Traganos
Frank
F
Darzynkiewicz
Zbigniew
Z
D015415:Biomarkers; D045744:Cell Line, Tumor; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D005434:Flow Cytometry / Q000379:methods*; D006657:Histones / Q000378:metabolism*; D006801:Humans; D010766:Phosphorylation / Q000187:drug effects; D016196:S Phase / Q000187:drug effects; D013194:Staining and Labeling; D019772:Topotecan / Q000494:pharmacology
D015415:Biomarkers; C468783:H2AX protein, human; D006657:Histones; D019772:Topotecan