Quantification of γ-H2AX Foci in Human Lymphocytes: A Method for Biological Dosimetry after Ionizing Radiation Exposure

Abstract

Recent studies have suggested that visualization of gamma-H2AX nuclear foci can be used to estimate exposure to very low doses of ionizing radiation. Although this approach is widely used for various purposes, its suitability for individual human biodosimetry has not yet been assessed. We therefore conducted such an assessment with the help of available software for observing and automatically scoring gamma-H2AX foci. The presence of gamma-H2AX foci was evaluated in human peripheral blood lymphocytes exposed ex vivo to gamma rays in a dose range of 0.02 to 2 Gy. We analyzed the response of gamma-H2AX to ionizing radiation in relation to dose, time after exposure, and individual variability. We constructed dose-effect calibration curves at 0.5, 8 and 16 h after exposure and evaluated the threshold of detection of the technique. The results show the promise of automatic gamma-H2AX scoring for a reliable assessment of radiation doses in a dose range of 0.6 Gy to 2 Gy up to 16 h after exposure. This gamma-H2AX-based assay may be useful for biodosimetry, especially for triage to distinguish promptly among individuals the ones who have received negligible doses from those with significantly exposures who are in need of immediate medical attention. However, additional in vivo experiments are needed for validation.

High throughput toxicity screening and intracellular detection of nanomaterials

“…Therefore, efforts have been made to improve this method, based on microscopy techniques such as imaging modalities in cell culture and in tissues,162 and computer‐assisted approaches 163. The computational approaches are supported by image analysis software such as NIH Image and its Windows counterpart ScionImage, ImageJ, HistoLab, AutoQuantX, or Image Pro, among others, which rely on different computational algorithms for foci identification or the calculation of quantitative foci parameters 164, 165, 166. High content imaging systems are also capable of γH2AX quantitation.…”

The second gamma-H2AX assay inter-comparison exercise carried out in the framework of the European biodosimetry network (RENEB)

“…Two or three laboratories received samples from the same donor. Based on previous experience using the five donors and on published studies (Roch-Lef evre et al 2010;Chua et al 2011), any variability between the donors would be expected to make only a small contribution to overall uncertainties.…”

“…The gamma-H2AX foci assay is a sensitive surrogate marker of radiation induced DNA double strand breaks (Ivashkevich et al 2012;Rothkamm et al 2015) and may be a useful early biodosimetry tool from hours to about three days post-exposure (Roch-Lef evre et al 2010;Mandina et al 2011). Several studies have shown that rapid processing time of the gamma-H2AX assay can produce dose estimates within a few hours of receiving a blood sample (Rothkamm et al 2013a;Ainsbury et al 2014).…”

Analysis of radioinduced DNA damages using Monte Carlo calculations at nanometric scale for different irradiation configurations

“…Radioinduced DSB in DNA can be localized within the cell nuclei by a fluorescence labeling of γ-H2AX around the damages [1]. Nevertheless, DSB are also produced during the cell cycle, mainly during the cell duplication.…”

“…The topological and quantitative analysis of these damages is now possible using immunofluorescence techniques [1]. Indeed, when a DSB occurs, many molecules of histone H2AX become phosphorilated (γ-H2AX) along megabase chromatin domains adjacent to the break as an early step of the DSB signaling and repair.…”