Extending the range of sensitivity for detecting DNA strand breaks

Several sensitive methods are now available to quantify the number of DNA strand breaks produced in mammalian cells by ionizing radiation. However, with one exception, these assays are sensitive to damage over a limited radiation dose range. The exception is the alkali-unwinding assay. This method is based on the premise that DNA strand breaks serve as points of unwinding when a cell is lysed and DNA denatured in dilute alkali (Ahnstrom & Erixon, 1973). The rate of strand separation is thus a function of the molecular weight of the DNA (Ahnstrom & Edvardsson, 1974). When the DNA solution is subsequently neutralized and sonicated, both single- and double-stranded DNA pieces are produced, with a greater proportion of DNA in single-stranded form corresponding to a greater number of strand breaks. Pieces of single- and double-stranded DNA can be easily separated using hydroxyapatite chromatography, and the percentage of DNA in double-stranded form is then a measure of DNA strand breakage. The advantages of this method include ease, accuracy and sensitivity for detecting DNA damage caused by low doses of ionizing radiation.