title: Techniques for gamma-H2AX detection.
abstract: When a double-strand break (DSB) forms in DNA, many molecules of histone H2AX present in the chromatin flanking the break site are rapidly phosphorylated. The phosphorylated derivative of H2AX is named gamma-H2AX, and the phosphorylation site is a conserved serine four residues from the C-terminus, 139 in mammals and 129 in budding yeast. An antibody to gamma-H2AX reveals that the molecules form a gamma-focus at the DSB site. The gamma-focus increases in size rapidly for 10-30 min after formation, and remains until the break is repaired. Studies have revealed that small numbers of gamma-foci are present in cells even without the purposeful introduction of DNA DSBs. These cryptogenic foci increase in number during senescence in culture and aging in mice. This chapter presents techniques for revealing gamma-H2AX foci in cultured cells, in metaphase spreads from cultured cells, in tissues, and in yeast.
authors.affiliation: Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, USA.
authors.forename: Asako
authors.initials: A
authors.lastname: Nakamura
authors.forename: Olga A
authors.initials: OA
authors.lastname: Sedelnikova
authors.forename: Christophe
authors.initials: C
authors.lastname: Redon
authors.forename: Duane R
authors.initials: DR
authors.lastname: Pilch
authors.forename: Natasha I
authors.initials: NI
authors.lastname: Sinogeeva
authors.forename: Robert
authors.initials: R
authors.lastname: Shroff
authors.forename: Michael
authors.initials: M
authors.lastname: Lichten
authors.forename: William M
authors.initials: WM
authors.lastname: Bonner
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D002478:Cells, Cultured; D004249:DNA Damage; D006657:Histones / Q000032:analysis* / Q000276:immunology; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D010766:Phosphorylation
chemical_list: D000906:Antibodies; D006657:Histones