VALIDATION OF CHROMOSOME PAINTING AS A BIODOSIMETER IN HUMAN PERIPHERAL LYMPHOCYTES FOLLOWING ACUTE EXPOSURE TO IONIZING-RADIATION IN-VITRO

Fluorescence in situ hybridization with chromosome-specific composite DNA probes ('chromosome painting') appears to be a useful tool for qua ntifying symmetrical cytogenetic damage. However, a thorough compariso n between chromosome painting and the conventional methods of GTG-band ing and dicentric analysis has not been performed. We have undertaken the validation of chromosome painting using human blood exposed in vit ro to Cs-137 gamma-rays at doses ranging from 0 to 400 cGy, then cultu red according to standard procedures and harvested at 52h. For paintin g, bound probes were detected either with fluoresceinated avidin and c ounterstained with propidium iodide, or with ChromoBlue WCP Probe and Giemsa. The first approach utilizes ultraviolet excitation in which pa inted chromosomes appear yellow and the remaining chromosomes appear r ed. The ChromoBlue labelling approach requires ordinary light microsco py in which painted chromosomes appear dark blue and the remaining chr omosomes appear light blue. With each method, exchanges between painte d and unpainted chromosomes appear bicoloured. Because only a fraction of all possible exchanges are detected, the number of metaphases exam ined is adjusted according to the fraction of the genome painted. We h ave performed painting by several methods, including fluorescence with chromosome 4 probe alone, fluorescence with probes for chromosomes 1, 3 and 4 simultaneously, and chromogenic painting with chromosome 4 pr obe alone. The results obtained by the various painting methods were c ompared with GTG-banded cells which were examined for both translocati ons and dicentrics. In addition, unbanded metaphases stained with Giem sa were scored for dicentrics. Our data show that the frequency of chr omosome exchanges detected by painting and banding agree with each oth er and with the number of dicentrics seen in unbanded cells, at least at doses of less-than-or-equal-to 200 cGy.