Elsevier

Radiation Measurements

Volume 42, Issues 6–7, July–August 2007, Pages 1143-1146
Radiation Measurements

Optimization of cytogenetic procedures for population triage in case of radiological emergency

https://doi.org/10.1016/j.radmeas.2007.05.044Get rights and content

Abstract

In case of accidental overexposure to ionizing radiation, the scoring of dicentrics in lymphocytes from blood is the current reference method to estimate the dose received. When only few individuals are accidentally overexposed, at least 500 cells are scored to have a good estimation of the dose. But such a practice is too time consuming when many people are exposed such as in a radiological emergency. In order to reduce the time required to estimate a dose, specific strategies have been developed in the laboratory.

Population triage based on the adaptation of the dicentrics assay: In order to reduce the analysis time it is possible to have a dose estimation based on only 50 cells analysed in an hour with a 95% confidence limit of 1 Gy.

Population triage based on the use of image analysis systems: The metaphase finder increases the speed of the scoring by a factor of 2. It is also possible to detect automatically the dicentrics. The system proposes to the operator some candidate dicentrics which are then verified manually. In this case, 50% of the dicentrics are correctly detected with a 95% confidence limit of 0.4 Gy.

Establishment of a network: Biological dosimetry laboratories are small, hence their capacity can be overloaded by a large number of suspected overexposed individuals. That is why international and national networks should be established, they should perform periodically intecomparisons and population triage exercises.

Population triage based on micronuclei assay: As micronuclei are easier to score than dicentrics, in an hour 500 binucleated cells can be scored resulting in a 95% confidence limit of about 0.5 Gy versus 1 Gy with the dicentric assay. However, the micronuclei technique is not efficient to estimate accurately the dose because it lacks specificity.

Use of γ-H2AX for population triage: This assay is based on the scoring of double-strand breaks. Whereas with the other technique a cell culture is required, here it is possible to estimate a dose 3 h after receiving the sample in the laboratory with a sensitivity below 0.5 Gy. But one major limitation is the decrease of the signal with time. Therefore, this assay can only be used in case of population triage starting within minutes post-exposure.

Introduction

Following exposure of cells to ionizing radiation, chromosomal aberrations are produced. The yield of unstable chromosomal aberrations as dicentrics and centric rings are used as dose indicators in biological dosimetry (IAEA, 2001). To have a statistically usable result 500 cells have to be scored to estimate the dose received. Scoring so many cells requires at least 2 days for one operator. This is not conceivable for a population triage where hundreds of persons may be exposed.

The aim of a population triage is to identify the victims who require early medical assistance from those less exposed. The population triage can be based on clinical signs such as the lymphocyte counts, nausea and diarrhoea, and local skin burns. However, those symptoms varies from one person to other and may result in false positive and false negative identifications. Biological indicator of dose could avoid such erroneous identifications. Such indicators should allow for a rapid assessment of a dose on a large number of victims.

Once the population has undergone early triage, it is required, in a second step, to better characterise the dose received. In this case, the time factor is less critical and a sensitive technique is necessary. Therefore, the ideal biological indicator of dose should be able to provide a rapid and sensitive response.

In a recent publication, the possibility to score the number of translocations detected using the fluorescent in situ hybridization technique has been evaluated for a population triage (Szeles et al., 2006). The reported result is a reduction of the sensitivity compared to the conventional dicentric approach.

Several other possibilities are proposed to deal with a large number of cases: (a) scoring only 50 cells using the dicentric assay; (b) the use of automation; (c) setting up national or international networks to be able to handle more samples; and (d) setting up new techniques that are faster and easier to carry out such as the micronuclei assay and the γ-H2AX assays. This paper discussed the advantages and disadvantages of these different approaches.

Section snippets

The dicentric assay: triage strategy

The standard technique (IAEA, 2001) has been adapted to improve the response time when triage is required to identify potentially irradiated individuals. The technique to prepare and analyse chromosomal aberrations is simplified and the number of metaphases observed is reduced. The observation of 50 cells requires 1 h scoring without the help of any image analysing system and allows a dose estimation with an uncertainty of ±1Gy (Voisin et al., 2001a, Voisin et al., 2001b). This margin of

Automatic dicentric detection

In order to reduce the time required for estimating the dose, image analysis systems were developed. Such systems, specific for biological dosimetry, are rare and not always commercialized. One of them is the METAFER developed by METASYSTEMS (Germany). It allows the automatic finding of metaphases, the acquisition of images at high magnification and the automatic detection of dicentrics stained uniformly with Giemsa (Schunck et al., 2004). The METAFER is the only system that allows the

Use of micronuclei in population triage

The interest of this technique is the simplicity in the scoring. Indeed, it is much easier to score micronuclei in binucleated cells than dicentrics among 46 chromosomes. This technique has already been described in Voisin et al. (2001a). A comparison of the main characteristics between the dicentric approach and the micronuclei one is presented in Table 1. The two techniques were compared during a triage exercise where 35 samples were exposed to doses from 0 to 3 Gy of 137cesium gamma

The use of γ-H2AX foci for a population triage

Another easy approach is the possibility to use immunostaining to detect double-strand breaks (DSBs). These lesions have been known as one of the most significant lesion producing lethal and mutagenic effects after exposure to ionising radiation. Recently, some inmunofluorescence studies have demonstrated that the number of radio-induced DNA DSBs correlates to the number of γ-H2AX nuclear foci that appear as the result of phosphorylation of H2AX histone (Brenner et al., 2003).

The advantage of

Conclusion

The biological triage is complementary to the clinical triage as some false positive and false negative can be identified. After comparison of several techniques, it appears that the automatic detection of dicentrics has the advantage to be fast and allows, in a second step, the manual scoring to better estimate the dose if necessary, however, it is not possible with this assay to discriminate heterogeneous exposures from whole body exposures. In the first step of triage situation the

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