An Optimized Method for Detecting Gamma-h2ax in Blood Cells Reveals a Significant Interindividual Variation in the Gamma-h2ax Response Among Humans.
From: Department of Clinical Chemistry and Transfusion Medicine, Sahlgrenska University Hospital, Göteborg University, SE-413 45 Göteborg, Sweden.
Nucleic acids research
- Publish Date: 2007
- ISSN: 1362-4962
- Volume: 35
- Issue: 5
- Pages: e36
- Medium: Internet
- Language: English
- Citation (JAMA): Ismail Ismail Hassan, Wadhra Tabasum Imran, Hammarsten Ola, et al. An Optimized Method for Detecting Gamma-h2ax in Blood Cells Reveals a Significant Interindividual Variation in the Gamma-h2ax Response Among Humans.. Nucleic Acids Res. 2007;35:e36
Abstract
Phosphorylation of histone H2AX on serine 139 (gamma-H2AX, gammaH2AX) occurs at sites flanking DNA double-strand breaks (DSBs) and can provide a measure of the number of DSBs within a cell. Here we describe a rapid and simple flow-cytometry-based method, optimized to measure gamma-H2AX in non-fixed peripheral blood cells. No DSB induced signal was observed in H2AX-/- cells indicating that our FACS method specifically recognized gamma-H2AX accumulation. The gamma-H2AX assay was capable of detecting DNA damage at levels 100-fold below the detection limit of the alkaline comet assay. The gamma-H2AX signal was quantitative with a linear increase of the gamma-H2AX signal over two orders of magnitude. We found that all nucleated blood cell types examined, including the short-lived neutrophils induce gamma-H2AX in response to DSBs. Interindividual difference in the gamma-H2AX signal in response to ionizing radiation and the DSB-inducing drug calicheamicin was almost 2-fold in blood cells from patients, indicating that the amount of gamma-H2AX produced in response to a given dose of radiation varies significantly in the human population. This simple method could be used to monitor response to radiation or DNA-damaging drugs.
Mesh Headings (Keywords): Aminoglycosides, Animals, Antibiotics, Antineoplastic, Cells, Cultured, DNA Breaks, Double-Stranded, Enediynes, Flow Cytometry, Fluorescein-5-isothiocyanate, Fluorescent Dyes, Histones, Humans, Leukocytes, Mice, Radiation, Ionizing
Check for Full Text / PubMed Unique Identifier (PMID): 17284459
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