Alternative methods for automatic dicentric detection in biological dosimetry

Roy L., Delbos M., Durand V., Sorokine-Durm I., Voisin P. The Second Euroconference on quantitative molecular cytogenetics, Salamanca, Spain, 26-28/04/2001

In case of accidental overexposure to ionizing radiation, the dicentrics produced in peripheral blood lymphocytes in metaphase, are used to estimate the radiation dose. However the number of dicentrics produced is low and up to 500 cells have to be scored to have a statistically significant estimation of the dose. Now manually it is possible to score up to 250 metaphases per cytogeneticist per day. Thus an automatic image analysis system has been developed to facilitate the scoring (IMSTAR, France). The first step of such a system is the automatic localisation of metaphases on the slide. The use of a metaphase finder has proved to reduce the time required to score a slide by a factor 2 to 4 according to the dose. But some functionalities are necessary to have such a gain benefit : (1) the adaptation of the metaphase detection parameters to each slide according to metaphases quality; (2) the possibility to test the metaphase detection parameters to verify their sensitivity and specificity; (3) the possibility to find metaphases both in bright and in fluorescence fields. When metaphases have been automatically located, it might be interesting to automate the dicentrics detection. One system allows the correct detection of 50% of the dicentrics (METASYSTEM, Germany). With homogeneous stained chromosomes it is difficult to reach a better efficiency as some chromosomes may present a constriction that is not necessarily a centromere. We therefore tried to detect dicentrics when centromeres are labelled by the hybridization in situ technique following by a fluorescent or an immunoperoxidase detection. The performances of chromosome detection are similar to those obtained with the conventional Giemsa stained chromosomes where all chromosomes are not always well separated. But then the centromeres can be easily localised by the presence of a spot. In fluorescence 95 % of the centromeres are correctly detected. Therefore it is possible to identify complete metaphases (i.d. 46 chromosomes) from the basis of the centromere number. Labels are used to identify dicentrics, normal chromosomes and fragments. Those labels can be more easily corrected than chromosome outlines therefore reducing the time of analysis. These results have to be compared to the results obtained with immunoperoxidase detection of centomeres.