Showing papers by "Joan Francesc Barquinero published in 2023"

Biodose Tools: An R shiny Application for Biological Dosimetry.

Abstract: Introduction In the event of a radiological accident or incident, the aim of biological dosimetry is to convert the yield of a specific biomarker of exposure to ionizing radiation into an absorbed dose. Since the 1980s, various tools have been used to deal with the statistical procedures needed for biological dosimetry, and in general those who made several calculations for different biomarkers were based on closed source software. Here we present a new open source program, Biodose Tools, that has been developed under the umbrella of RENEB (Running the European Network of Biological and retrospective Physical dosimetry).Material and Methods The application has been developed using the R programming language and the shiny package as a framework to create a user-friendly online solution. Since no unique method exists for the different mathematical processes, several meetings and periodic correspondence were held in order to reach a consensus on the solutions to be implemented.Results The current version 3.6.1 supports dose-effect fitting for dicentric and translocation assay. For dose estimation Biodose Tools implements those methods indicated in international guidelines and a specific method to assess heterogeneous exposures. The app can include information on the irradiation conditions to generate the calibration curve. Also, in the dose estimate, information about the accident can be included as well as the explanation of the results obtained. Because the app allows generating a report in various formats, it allows traceability of each biological dosimetry study carried out. The app has been used globally in different exercises and training, which has made it possible to find errors and improve the app itself. There are some features that still need consensus, such as curve fitting and dose estimation using micronucleus analysis. It is also planned to include a package dedicated to interlaboratory comparisons and the incorporation of Bayesian methods for dose estimation.Conclusion Biodose Tools provides an open-source solution for biological dosimetry laboratories. The consensus reached helps to harmonize the way in which uncertainties are calculated. In addition, because each laboratory can download and customize the app's source code, it offers a platform to integrate new features.

RENEB Inter-Laboratory Comparison 2021: The Dicentric Chromosome Assay

Abstract: After large-scale radiation accidents where many individuals are suspected to be exposed to ionizing radiation, biological and physical retrospective dosimetry assays are important tools to aid clinical decision making by categorizing individuals into unexposed/minimally, moderately or highly exposed groups. Quality-controlled inter-laboratory comparisons of simulated accident scenarios are regularly performed in the frame of the European legal association RENEB (Running the European Network of Biological and Physical retrospective Dosimetry) to optimize international networking and emergency readiness in case of large-scale radiation events. In total 33 laboratories from 22 countries around the world participated in the current RENEB inter-laboratory comparison 2021 for the dicentric chromosome assay. Blood was irradiated in vitro with X rays (240 kVp, 13 mA, ∼75 keV, 1 Gy/min) to simulate an acute, homogeneous whole-body exposure. Three blood samples (no. 1: 0 Gy, no. 2: 1.2 Gy, no. 3: 3.5 Gy) were sent to each participant and the task was to culture samples, to prepare slides and to assess radiation doses based on the observed dicentric yields from 50 manually or 150 semi-automatically scored metaphases (triage mode scoring). Approximately two-thirds of the participants applied calibration curves from irradiations with γ rays and about 1/3 from irradiations with X rays with varying energies. The categorization of the samples in clinically relevant groups corresponding to individuals that were unexposed/minimally (0–1 Gy), moderately (1–2 Gy) or highly exposed (>2 Gy) was successfully performed by all participants for sample no. 1 and no. 3 and by ≥74% for sample no. 2. However, while most participants estimated a dose of exactly 0 Gy for the sham-irradiated sample, the precise dose estimates of the samples irradiated with doses >0 Gy were systematically higher than the corresponding reference doses and showed a median deviation of 0.5 Gy (sample no. 2) and 0.95 Gy (sample no. 3) for manual scoring. By converting doses estimated based on γ-ray calibration curves to X-ray doses of a comparable mean photon energy as used in this exercise, the median deviation decreased to 0.27 Gy (sample no. 2) and 0.6 Gy (sample no. 3). The main aim of biological dosimetry in the case of a large-scale event is the categorization of individuals into clinically relevant groups, to aid clinical decision making. This task was successfully performed by all participants for the 0 Gy and 3.5 Gy samples and by 74% (manual scoring) and 80% (semiautomatic scoring) for the 1.2 Gy sample. Due to the accuracy of the dicentric chromosome assay and the high number of participating laboratories, a systematic shift of the dose estimates could be revealed. Differences in radiation quality (X ray vs. γ ray) between the test samples and the applied dose effect curves can partly explain the systematic shift. There might be several additional reasons for the observed bias (e.g., donor effects, transport, experimental conditions or the irradiation setup) and the analysis of these reasons provides great opportunities for future research. The participation of laboratories from countries around the world gave the opportunity to compare the results on an international level.

RENEB Inter-Laboratory Comparison 2021: Inter-Assay Comparison of Eight Dosimetry Assays

Abstract: Tools for radiation exposure reconstruction are required to support the medical management of radiation victims in radiological or nuclear incidents. Different biological and physical dosimetry assays can be used for various exposure scenarios to estimate the dose of ionizing radiation a person has absorbed. Regular validation of the techniques through inter-laboratory comparisons (ILC) is essential to guarantee high quality results. In the current RENEB inter-laboratory comparison, the performance quality of established cytogenetic assays [dicentric chromosome assay (DCA), cytokinesis-block micronucleus assay (CBMN), stable chromosomal translocation assay (FISH) and premature chromosome condensation assay (PCC)] was tested in comparison to molecular biological assays [gamma-H2AX foci (gH2AX), gene expression (GE)] and physical dosimetry-based assays [electron paramagnetic resonance (EPR), optically or thermally stimulated luminescence (LUM)]. Three blinded coded samples (e.g., blood, enamel or mobiles) were exposed to 0, 1.2 or 3.5 Gy X-ray reference doses (240 kVp, 1 Gy/min). These doses roughly correspond to clinically relevant groups of unexposed to low exposed (0–1 Gy), moderately exposed (1–2 Gy, no severe acute health effects expected) and highly exposed individuals (>2 Gy, requiring early intensive medical care). In the frame of the current RENEB inter-laboratory comparison, samples were sent to 86 specialized teams in 46 organizations from 27 nations for dose estimation and identification of three clinically relevant groups. The time for sending early crude reports and more precise reports was documented for each laboratory and assay where possible. The quality of dose estimates was analyzed with three different levels of granularity, 1. by calculating the frequency of correctly reported clinically relevant dose categories, 2. by determining the number of dose estimates within the uncertainty intervals recommended for triage dosimetry (±0.5 Gy or ±1.0 Gy for doses <2.5 Gy or >2.5 Gy), and 3. by calculating the absolute difference (AD) of estimated doses relative to the reference doses. In total, 554 dose estimates were submitted within the 6-week period given before the exercise was closed. For samples processed with the highest priority, earliest dose estimates/categories were reported within 5–10 h of receipt for GE, gH2AX, LUM, EPR, 2–3 days for DCA, CBMN and within 6–7 days for the FISH assay. For the unirradiated control sample, the categorization in the correct clinically relevant group (0–1 Gy) as well as the allocation to the triage uncertainty interval was, with the exception of a few outliers, successfully performed for all assays. For the 3.5 Gy sample the percentage of correct classifications to the clinically relevant group (≥2 Gy) was between 89–100% for all assays, with the exception of gH2AX. For the 1.2 Gy sample, an exact allocation to the clinically relevant group was more difficult and 0–50% or 0–48% of the estimates were wrongly classified into the lowest or highest dose categories, respectively. For the irradiated samples, the correct allocation to the triage uncertainty intervals varied considerably between assays for the 1.2 Gy (29–76%) and 3.5 Gy (17–100%) samples. While a systematic shift towards higher doses was observed for the cytogenetic-based assays, extreme outliers exceeding the reference doses 2–6 fold were observed for EPR, FISH and GE assays. These outliers were related to a particular material examined (tooth enamel for EPR assay, reported as kerma in enamel, but when converted into the proper quantity, i.e. to kerma in air, expected dose estimates could be recalculated in most cases), the level of experience of the teams (FISH) and methodological uncertainties (GE). This was the first RENEB ILC where everything, from blood sampling to irradiation and shipment of the samples, was organized and realized at the same institution, for several biological and physical retrospective dosimetry assays. Almost all assays appeared comparably applicable for the identification of unexposed and highly exposed individuals and the allocation of medical relevant groups, with the latter requiring medical support for the acute radiation scenario simulated in this exercise. However, extreme outliers or a systematic shift of dose estimates have been observed for some assays. Possible reasons will be discussed in the assay specific papers of this special issue. In summary, this ILC clearly demonstrates the need to conduct regular exercises to identify research needs, but also to identify technical problems and to optimize the design of future ILCs.

RENEB Inter-Laboratory Comparison 2021: The FISH-Based Translocation Assay

Abstract: Translocation analysis using fluorescence in situ hybridization (FISH) is the method of choice for dose assessment in case of chronic or past exposures to ionizing radiation. Although it is a widespread technique, unlike dicentrics, the number of FISH-based inter-laboratory comparisons is small. For this reason, although the current Running the European Network of Biological and Physical retrospective Dosimetry (RENEB) inter-laboratory comparison 2021 was designed as a fast response to a real emergency scenario, it was considered a good opportunity to perform an inter-laboratory comparison using the FISH technique to gain further experience. The Bundeswehr Institute of Radiobiology provided peripheral blood samples from one healthy human volunteer. Three test samples were irradiated with blinded doses of 0, 1.2, and 3.5 Gy, respectively. Samples were then sent to the seven participating laboratories. The FISH technique was applied according to the standard procedure of each laboratory. Both, the frequency of translocations and the estimated dose for each sample were sent to the coordinator using a special scoring sheet for FISH. All participants sent their results in due time. However, although it was initially requested to send the results based on the full analysis, evaluating 500 equivalent cells, most laboratories only sent the results based on triage, with a smaller number of analyzed cells. In the triage analysis, there was great heterogeneity in the number of equivalent cells scored. On the contrary, for the full analysis, this number was more homogeneous. For all three samples, one laboratory showed outlier yields compared to the other laboratories. Excluding these results, in the triage analysis, the frequency of translocations in sample no. 1 ranged from 0 to 0.013 translocations per cell, and for samples no. 2 and no. 3 the genomic mean frequency were 0.27 ± 0.03 and 1.47 ± 0.14, with a coefficient of variation of 0.29 and 0.23 respectively. Considering only results obtained in the triage analysis for sample no. 1, all laboratories, except one, classified this sample as the non-irradiated one. For sample no. 2, excluding the outlier value, the mean reported dose was 1.74 ± 0.16 Gy indicating a mean deviation of about 0.5 Gy to the delivered dose of 1.2 Gy. For sample no. 3 the mean dose estimated was 4.21 ± 0.21 Gy indicating a mean deviation of about 0.7 Gy to the delivered dose of 3.5 Gy. In the frame of RENEB, this is the second FISH-based inter-laboratory comparison. The whole exercise was planned as a response to an emergency, therefore, a triage analysis was requested for all the biomarkers except for FISH. Although a full analysis was initially requested for FISH, most of the laboratories reported only a triage-based result. The main reason is that it was not clearly stated what was required before starting the exercise. Results show that most of the laboratories successfully discriminated unexposed and irradiated samples from each other without any overlap. A good agreement in the observed frequencies of translocations was observed but there was a tendency to overestimate the delivered doses. Efforts to improve the harmonization of this technique and subsequent exercises to elucidate the reason for this trend should be promoted.

What We Have Learned from RENEB Inter-Laboratory Comparisons Since 2012 With Focus on ILC 2021

Abstract: Inter-laboratory exercises are important tools within the European network for biological dosimetry and physical retrospective dosimetry (RENEB) to validate and improve the performance of member laboratories and to ensure an operational network with high quality standards for dose estimations in case of a large-scale radiological or nuclear event. In addition to the RENEB inter-laboratory comparison 2021, several inter-laboratory comparisons have been performed in the frame of RENEB for a number of assays in recent years. This publication gives an overview of RENEB inter-laboratory comparisons for biological dosimetry assays in the past and a final summary of the challenges and lessons learnt from the RENEB inter-laboratory comparison 2021. In addition, the dose estimates of all RENEB inter-laboratory comparisons since 2013 that have been conducted for the dicentric chromosome assay, the most established and applied assay, are compared and discussed.

Use of a plasma focus device to study pulsed x-ray effects on peripheral blood lymphocytes: Analysis of chromosome aberrations

Abstract: X-ray pulses (full width at half maximum ∼ 90 ns, dose rate ∼ 107 Gy s−1) were used to irradiate the monolayer of peripheral blood mononucleated cells using the PF-2kJ kilojoule plasma focus device. Four different exposure conditions were evaluated using 5, 10, 20, and 40 pulses, with the mean dose measured by TLD-100 being 0.12 ± 0.02 mGy, 0.14 ± 0.03 mGy, 0.22 ± 0.06 mGy, and 0.47 ± 0.09 mGy, respectively. Cytogenetic analysis showed an increase in all types of chromosomal aberrations following exposure to x-ray pulses. The distribution of dicentrics and centric rings was overdispersed after 5, 10, 20, and 40 pulses. Additionally, after 20 and 40 pulses, the presence of tricentric chromosomes is detected. Chromosome aberration frequencies found in this study were always higher than the estimated frequencies of chromosome aberrations using published dose–effect curves for conventional radiation sources. The overdispersion observed, the elevated maximum relative biological effectiveness (RBEM) and the presence of tricentric chromosomes at the relatively low doses of exposure (<0.5 Gy) seem to indicate that low doses of pulsed x-rays of low energy show similar biological effects as those observed for high-LET radiation. X-ray pulses emitted by PF-2kJ were found to be more efficient in inducing chromosome aberrations, even more than α particles.

International comparison exercise for biological dosimetry after exposures with neutrons performed at two irradiation facilities as part of the BALANCE project.

Abstract: In the case of a radiological or nuclear event, biological dosimetry can be an important tool to support clinical decision-making. During a nuclear event, individuals might be exposed to a mixed field of neutrons and photons. The composition of the field and the neutron energy spectrum influence the degree of damage to the chromosomes. During the transatlantic BALANCE project, an exposure similar to a Hiroshima-like device at a distance of 1.5 km from the epicenter was simulated and biological dosimetry based on dicentric chromosomes was performed to evaluate the participants ability to discover unknown doses and to test the influence of differences in neutron spectra. In a first step, calibration curves were established by irradiating blood samples with 5 doses in the range of 0 Gy to 4 Gy at two different facilities in Germany (PTB) and USA (CINF). The samples were sent to eight participating laboratories from the RENEB network and dicentric chromosomes were scored by each participant. Next, blood samples were irradiated with 4 blind doses in each of the two facilities and sent to the participants to provide dose estimates based on the established calibration curves. Manual and semi-automatic scoring of dicentric chromosomes were evaluated for their applicability to neutron exposures. Moreover, the biological effectiveness of the neutrons from the two irradiation facilities was compared. The calibration curves from samples irradiated at CINF showed a 1.4 times higher biological effectiveness compared to samples irradiated at PTB. For manual scoring of dicentric chromosomes, the doses of the test samples were mostly successfully resolved based on the calibration curves established during the project. For semi-automatic scoring, the dose estimation for the test samples was less successful. Doses >2 Gy in the calibration curves revealed non-linear associations between dose and dispersion index of the dicentric counts, especially for manual scoring. The differences in the biological effectiveness between the irradiation facilities suggested that the neutron energy spectrum can have a strong impact on the dicentric counts.

Evaluation of γ-H2AX foci distribution among different peripheral blood mononucleated cell subtypes.

Abstract: INTRODUCTION The detection of γ-H2AX foci in peripheral blood mononucleated cells (PBMCs) has been incorporated as an early assay for biological dosimetry. However, overdispersion in the γ-H2AX foci distribution is generally reported. In a previous study from our group, it was suggested that overdispersion could be caused by the fact that when evaluating PBMCs, different cell subtypes are analyzed, and that these could differ in their radiosensitivity. This would cause a mixture of different frequencies that would result in the overdispersion observed. OBJECTIVES The objective of this study was to evaluate both the possible differences in the radiosensitivities of the different cell subtypes present in the PBMCs and to evaluate the distribution of γ-H2AX foci in each cell subtype. MATERIALS AND METHODS Peripheral blood samples from three healthy donors were obtained and total PBMCs, and CD3+, CD4+, CD8+, CD19+, and CD56+ cells were separated. Cells were irradiated with 1 and 2 Gy and incubated at 37 °C for 1, 2, 4, and 24 h. Sham-irradiated cells were also analyzed. γ-H2AX foci were detected after immunofluorescence staining and analyzed automatically using a Metafer Scanning System. For each condition, 250 nuclei were considered. RESULTS When the results from each donor were compared, no observable significant differences between donors were observed. When the different cell subtypes were compared, CD8+ cells showed the highest mean of γ-H2AX foci in all post-irradiation time points. The cell type that showed the lowest γ-H2AX foci frequency was CD56+. The frequencies observed in CD4+ and CD19+ cells fluctuated between CD8+ and CD56+ without any clear pattern. For all cell types evaluated, and at all post-irradiation times, overdispersion in γ-H2AX foci distribution was significant. Independent of the cell type evaluated the value of the variance was four times greater than that of the mean. CONCLUSION Although different PBMC subsets studied showed different radiation sensitivity, these differences did not explain the overdispersion observed in the γ-H2AX foci distribution after exposure to IR.