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?

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 24. h 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 4. Gy, acute simulated partial body (4. Gy to 50% of cells) and protracted exposures (4. Gy over 24. h) were analysed. Data from all laboratories could be satisfactorily fitted with linear dose response functions. Average yields observed at 4. h post exposure were 2-4 times higher than at 24. h 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.