RENEB Inter-Laboratory Comparison 2021: The Gamma-H2AX Foci Assay.

The Running the European Network of biological and retrospective dosimetry (RENEB) network of laboratories has a range of biological and physical dosimetry assays that can be deployed in the event of a radiation incident to provide exposure assessment. To maintain operational capability and provide training, RENEB runs regular inter-laboratory comparison (ILC) exercises. The RENEB ILC2021 was carried out with all the biological and physical dosimetry assays employed in the network. The focus of this paper is to evaluate the results from 6 laboratories that took part using the gamma-H2AX radiation-induced foci assay. For two laboratories this was their first RENEB ILC. Blood samples were homogenously exposed to 240 kVp X rays (1 Gy/min) to provide calibration data, (0-4 Gy), and a few weeks later three blind coded test samples, (0, 1.2 and 3.5 Gy) were prepared. All samples were allowed a 2 h repair time at 37°C before being transported, on ice packs, to the participating laboratories. On arrival, the samples were processed, scored either manually or automatically for gamma-H2AX foci and dose estimates for the 3 blind coded samples sent to the organizing laboratory. The temperature of samples during transit and the time taken to report the dose estimates were recorded. Subsequent examination of the data from each laboratory used the doses estimates to assign triage categories to the samples. After receipt of the samples, the quickest report of dose estimates was 4.6 h. Analysis of variance revealed that the laboratory carrying out the assay had a significant effect on the foci yield (P < 0.001) for the calibration data, but not on the dose estimates of the blind coded samples (P = 0.101). All laboratories correctly identified the unirradiated and irradiated samples, although the dose estimates for the latter tended to under-estimate the dose. Two participants seriously under-estimated the dose for the highly exposed sample, which resulted in the sample being placed in the lowest triage category not the highest. However, this under-estimation resulted from the samples not remaining cold during shipment, due to a delay in transit and was not related to the experience of the participating laboratory. Overall, the RENEB network laboratories have demonstrated it is possible to quickly identify a recent whole-body acute exposure using the gamma-H2AX assay within the conditions of the ILC. In addition, an ILC provides a useful training and harmonization exercise for laboratories.

Effect of curcumin on γ-ray-induced cell response.

The purpose of the present study is to evaluate the effect of curcumin as a natural compound against radiation induced γ-foci and stable chromosome aberrations. Whole blood samples form three human volunteers were pretreated with curcumin at different concentrations (0.5, 10, 20 and 100 µg/ml). After 1-hour incubation, the lymphocytes were exposed to γ-rays (0.05, 0.5, 1 and 2 Gy). Radiation induced changes in cells were quantified using γ-H2AX/53BP1 assay and FISH analysis. Our results have shown that curcumin significantly reduced the frequency of both γ-foci and translocations. We found concentration-dependent increase of curcumin protective effect on γ-H2AX/53BP1 foci formation at all radiation doses. Concerning the translocations, after 0.05 and 0.5 Gy γ-rays the values of genomic frequencies are comparable within each dose and we did not observe any impact of curcumin. The most protective effect after 1 Gy exposure was found at 100 µg/ml curcumin. At 2 Gy irradiation, the maximum protection was achieved at 0.5 and 10 µg/ml of curcumin. Concentrations of 20 and 100 µg/ml also prevent lymphocytes but to less extent. Our in vitro study indicates radioprotective efficacy of curcumin against γ-ray induced damages in human lymphocytes. This observation suggests that curcumin may play a role to protect patients undergoing radiological procedures.

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

PURPOSE: Within the EU RENEB project, seven laboratories have taken part in training and harmonisation activities to strengthen triage gamma-H2AX-based radiation exposure assessment. This has culminated in a second triage biodosimetry exercise. MATERIALS AND METHODS: Whole blood and separated lymphocyte samples were homogenously irradiated with 60Co gamma rays at 0.5, 2.5 (blind samples), 0 and 2 Gy (reference samples). Following post-exposure incubations of 4 and 24 h, 16 samples were shipped on ice packs to each partner. The samples were stained and scored for gamma-H2AX foci, using manual and/or automated fluorescence microscope scoring strategies. Dose estimates were obtained and used to assign triage categories to the samples. RESULTS: Average dose estimates across all the laboratories correlated well with true doses. The most accurate assignment of triage category was achieved by manual scoring of the 4-h blood and lymphocyte samples. Only three samples out of a total of 46 were miscategorized in a way that could have adversely effected the clinical management of a radiation casualty. CONCLUSIONS: This inter-comparison exercise has demonstrated that following a recent acute radiation exposure, the gamma-H2AX assay could be a useful triage tool that can be successfully applied across a network of laboratories.

Chromosome analysis of nuclear power plant workers using fluorescence in situ hybridization and Giemsa assay.

The aim of this study was to evaluate the genotoxic effects of ionizing radiation in vivo in exposed Bulgarian nuclear power plant workers by using classical cytogenetic and molecular cytogenetic analyses of peripheral lymphocytes. Chromosome analysis using fluorescence in situ hybrydization (FISH) and Giemsa techniques was undertaken on 63 workers and 45 administrative staff controls from the Bulgarian Nuclear Power Plant. Using the Giemsa method, the frequencies of cells studied with chromosome aberrations, dicentrics plus rings and chromosome fragments in the radiation workers were significantly higher compared with the control group (P = 0.044, P = 0.014, and P = 0.033, respectively). A significant association between frequencies of dicentrics plus rings and accumulated doses was registered (P < 0.01). In the present study, a FISH cocktail of whole chromosome paints for chromosomes 1, 4 and 11 was used. A significant association between frequency of translocations and accumulated doses was also observed (P < 0.001). Within the control group, a correlation was found between age and the spontaneous frequency of translocations. No correlation was found between smoking status and frequency of translocations. When compared with the control group, workers with accumulated doses up to 100 mSv showed no increase in genome translocation frequency, whereas workers with accumulated doses from 101 to 200 mSv showed a statistically significant doubling of genome translocation frequency (P = 0.009). Thus, in cases of chronic exposure and for purposes of retrospective dosimetry, the genome frequency of translocations is a more useful marker for evaluation of genotoxic effects than dicentric frequency.

The use of the dicentric assay for biological dosimetry for radiation accidents in Bulgaria.

This paper details the construction of a 137Cs gamma calibration curve that has been established for dicentric assay and the testing and validation of the curve through biological dosimetry in three situations of suspected workplace overexposure that arose accidentally or through negligence or lack of appropriate safety measures. The three situations were: (1) suspected 137Cs contamination in a factory air supply; (2) suspected exposure to an industrial 192Ir source; and (3) accidental exposure of construction workers to radiation from a 60Co radiotherapy source in a hospital medical physics department. From a total of 24 potentially-exposed subjects, only one worker was found to have a statistically significant dose (0.16 Gy, 95% confidence intervals 0.02-0.43 Gy). In all other cases, the main function of the biological dosimetry was to reassure the subjects that any dose received was low.