Rapid biodosimetry tools are required to assist with triage in the case of a large-scale radiation incident. Here, we aimed to determine the dose-assessment accuracy of the well-established dicentric chromosome assay (DCA) and cytokinesis-block micronucleus assay (CBMN) in comparison to the emerging γ-H2AX foci and gene expression assays for triage mode biodosimetry and radiation injury assessment. Coded blood samples exposed to 10 X-ray doses (240 kVp, 1 Gy/min) of up to 6.4 Gy were sent to participants for dose estimation. Report times were documented for each laboratory and assay. The mean absolute difference (MAD) of estimated doses relative to the true doses was calculated. We also merged doses into binary dose categories of clinical relevance and examined accuracy, sensitivity and specificity of the assays. Dose estimates were reported by the first laboratories within 0.3–0.4 days of receipt of samples for the γ-H2AX and gene expression assays compared to 2.4 and 4 days for the DCA and CBMN assays, respectively. Irrespective of the assay we found a 2.5–4-fold variation of interlaboratory accuracy per assay and lowest MAD values for the DCA assay (0.16 Gy) followed by CBMN (0.34 Gy), gene expression (0.34 Gy) and γ-H2AX (0.45 Gy) foci assay. Binary categories of dose estimates could be discriminated with equal efficiency for all assays, but at doses ≥1.5 Gy a 10% decrease in efficiency was observed for the foci assay, which was still comparable to the CBMN assay. In conclusion, the DCA has been confirmed as the gold standard biodosimetry method, but in situations where speed and throughput are more important than ultimate accuracy, the emerging rapid molecular assays have the potential to become useful triage tools.

In case of a large-scale radiological incident, the pooling of ressources by networks can enhance the rapid classification of individuals in medically relevant treatment groups based on the DCA. The performance of the RENEB network as a whole has clearly benefited from harmonization processes and specific training activities for the network partners.

The focus of the study is an intercomparison of laboratories' dose-assessment performances using the γ-H2AX foci assay as a diagnostic triage tool for rapid individual radiation dose assessment. Homogenously X-irradiated (240 kVp, 1 Gy/min) blood samples for establishing calibration data (0.25-4 Gy) as well as blinded test samples (0.1-6.4 Gy) were incubated at 37°C for 2 and 24 h (repair time) and sent to the participants. The foci assay was performed according to protocols individually established in participating laboratories and therefore varied. The time taken to report dose estimates was documented for each laboratory. Additional information concerning laboratory organization/characteristics as well as assay performance was collected. The mean absolute difference (MAD) of estimated doses relative to the actual doses was calculated and radiation doses were merged into four triage categories reflecting clinical relevance to calculate accuracy, sensitivity and specificity. First γ-H2AX based dose estimates were reported 7 h after sample receipt. Estimates were similarly accurate for 2 and 24 h repair times, providing scope for its use in the early phase of a radiation exposure incident. Equal accuracy was achieved by scoring 20, 30, 40 or 50 cells per sample. However, MAD values of 0.5-0.7 Gy and 1.3-1.7 Gy divided the data sets into two groups, driven mainly by the considerable differences in foci yields between calibration and blind samples. Foci yields also varied dramatically between laboratories, highlighting reproducibility issues as an important caveat of the foci assay. Nonetheless, foci counts could distinguish high- and low-dose samples in all data sets and binary dose categories of clinical significance could be discriminated with satisfactory accuracy (mean 84%, ±0.03 SEM). Overall, the results suggest that the γ-H2AX assay is a useful tool for rapidly screening individuals for significant exposures that occurred up to at least 24 h earlier, and may help to prioritize cytogenetic dosimetry follow-up.

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Ataxia-telangiectasia (A-T) is an autosomal recessive neurodegenerative disorder characterized by radiosensitivity, genomic instability, and predisposition to cancer. A-T is caused by biallelic mutations in the ataxia-telangiectasia mutated (ATM) gene, but heterozygous carriers, though apparently healthy, are believed to be at increased risk for cancer and more sensitive to ionizing radiation than the general population. Despite progress in functional and sequencing-based assays, no straightforward, rapid, and inexpensive test is available for the identification of A-T homozygotes and heterozygotes, which is essential for diagnosis, genetic counseling, and carrier prediction. The oncosuppressor p53 prevents genomic instability and centrosomal amplification. During mitosis, p53 localizes at the centrosome in an ATM-dependent manner. We capitalized on the latter finding and established a simple, fast, minimally invasive, reliable, and inexpensive test to determine mutant ATM zygosity. The percentage of mitotic lymphoblasts or PBMCs bearing p53 centrosomal localization clearly discriminated among healthy donors (>75%), A-T heterozygotes (40%-56%), and A-T homozygotes (<30%). The test is specific for A-T, independent of the type of ATM mutations, and recognized tumor-associated ATM polymorphisms. In a preliminary study, our test confirmed that ATM is a breast cancer susceptibility gene. These data open the possibility of cost-effective, early diagnosis of A-T homozygotes and large-scale screenings for heterozygotes.