The use of the same image analysis system to detect genetic damage in human lymphocytes treated with doxorubicin in the Comet and fluoresence in situ hybridisation (FISH) assays

Abstract

Two assays, the alkaline single cell gel electrophoresis (Comet) assay and the fluorescence in situ hybridisation (FISH) of a whole chromosome or `chromosome painting' assay have gained importance in recent years as witnessed by the increasing yield of scientific literature using these techniques. Thus, it would be useful to have one system to measure both endpoints. In the present communication, a cost-effective electronic imaging system developed by Kinetic Imaging Ltd., UK, has been used to measure, after treatment of human lymphocytes with doxorubicin, DNA damage in the Comet assay (using software package KOMET) and chromosome damage with the FISH technique (using software package KROMASCAN). The chromosome damage has been detected using chromoprobe™-M for chromosome 1 and compared with chromosome damage measured using the conventional Giemsa staining technique. In all three assays, cycling cells were treated, after phytohaemagglutinin stimulation, at 48 h for about 20 h, which resulted in statistically significant dose-related responses in each assay. In non-cycling cells there was no increase in damage in the Comet assay, but there was in the chromosome assays. The FISH assay was only conducted in cycling cells, since the probe used was metaphase-specific. At the highest doses of doxorubicin used, FISH and conventional chromosome assays had similar sensitivities.