DNA damage detection technique applying time-resolved fluorescence measurements

A novel DNA damage detection technique based on the characteristic fluorescence lifetimes exhibited by Pico-Green-single-stranded DNA and -double-stranded DNA complexes is employed to establish the damage produced on DNA isolated from sheep white blood cells following gamma radiation. This technique, which incorporates key concepts such as alkaline unwinding buffers and higher unwinding rates of damaged DNA compared to undamaged DNA, allows for the differentiation of DNA damage resulting from doses of gamma radiation in the 0-100-Gy range, with the potential of analyzing samples consisting of as little as 10(4) cells. Experiments were carried out using commercial DNA sources as well as DNA isolated from sheep white blood cells, suggesting its potential for use with isolated DNA from virtually any eukaryotic cell.