Damage to cellular DNA is implicated in the early stages of carcinogenesis and in the cytotoxicity of many anticancer agents, including ionizing radiation. CE is a sensitive technique for measuring cellular levels of DNA damage through the detection of specific DNA lesions by specific monoclonal antibodies.
The detection is done by the use of a secondary antibody labeled with a fluorochrome. This method has been applied to the determination of thymine glycol residues in DNA generated by irradiation of cells during radiation therapy. The sensitivity of detection is in the range of zeptomoles.
Another way to detect DNA damage is the evaluation of DNA laddering during apoptosis. A key step in the onset of apoptosis is cleavage of the genomic DNA between nucleosomes, resulting in polynucleosome-sized fragments of DNA that give rise to a characteristic DNA laddering pattern.
A fluorescent intercalating dye is used to label the DNA molecules that will be resolved by CGE. Also, the use of a DNA standard curve allows quantification of the apoptotic DNA. The use of CGE with LIF detection permits analysis of DNA laddering with improved automation and much greater sensitivity.
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