title: Manual versus automated γ-H2AX foci analysis across five European laboratories: can this assay be used for rapid biodosimetry in a large scale radiation accident?
abstract: The identification of severely exposed individuals and reassurance of the 'worried well' are of prime importance for initial triage following a large scale radiation accident. We aim to develop the γ-H2AX foci assay into a rapid biomarker tool for use in accidents. Here, five laboratories established a standard operating procedure and analysed 100 ex vivo γ-irradiated, 4 or 24h incubated and overnight-shipped lymphocyte samples from four donors to generate γ-H2AX reference data, using manual and/or automated foci scoring strategies. In addition to acute, homogeneous exposures to 0, 1, 2 and 4Gy, acute simulated partial body (4Gy to 50% of cells) and protracted exposures (4Gy over 24h) were analysed. Data from all laboratories could be satisfactorily fitted with linear dose response functions. Average yields observed at 4h post exposure were 2-4 times higher than at 24h and varied considerably between laboratories. Automated scoring caused larger uncertainties than manual scoring and was unable to identify partial exposures, which were detectable in manually scored samples due to their overdispersed foci distributions. Protracted exposures were detectable but doses could not be accurately estimated with the γ-H2AX assay. We conclude that the γ-H2AX assay may be useful for rapid triage following a recent acute radiation exposure. The potentially higher speed and convenience of automated relative to manual foci scoring needs to be balanced against its compromised accuracy and inability to detect partial body exposures. Regular re-calibration or inclusion of reference samples may be necessary to ensure consistent results between laboratories or over long time periods.
authors: Public Health England, Centre for Radiation, Chemical and Environmental Hazards, Chilton, Didcot, Oxon OX11 0RQ, UK. Electronic address: kai.rothkamm@phe.gov.uk.
authors: Kai
authors: K
authors: Rothkamm
authors: Stephen
authors: S
authors: Barnard
authors: Elizabeth A
authors: EA
authors: Ainsbury
authors: Jenna
authors: J
authors: Al-Hafidh
authors: Joan-Francesc
authors: JF
authors: Barquinero
authors: Carita
authors: C
authors: Lindholm
authors: Jayne
authors: J
authors: Moquet
authors: Marjo
authors: M
authors: Perälä
authors: Sandrine
authors: S
authors: Roch-Lefèvre
authors: Harry
authors: H
authors: Scherthan
authors: Hubert
authors: H
authors: Thierens
authors: Anne
authors: A
authors: Vral
authors: Veerle
authors: V
authors: Vandersickel
mesh_terms: D001331:Automation; D004307:Dose-Response Relationship, Radiation; D005060:Europe; D005720:Gamma Rays / Q000009:adverse effects*; D006657:Histones / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D007753:Laboratories / Q000592:standards*; D008214:Lymphocytes / Q000378:metabolism* / Q000528:radiation effects; D008856:Microscopy, Fluorescence; D011834:Radiation Monitoring / Q000379:methods*; D018788:Radioactive Hazard Release / Q000517:prevention & control*; D013997:Time Factors
chemical_list: C468783:H2AX protein, human; D006657:Histones
keywords: Biological dosimetry; Biomarker; DNA double-strand break; Gamma-H2AX; Ionizing radiation; Radiation-induced foci