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Single cell gel electrophoresis: Detection of DNA damage at different levels of sensitivity
Author(s) -
Angelis Karel J.,
Dušinská Mária,
Collins Andrew R.
Publication year - 1999
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/(sici)1522-2683(19990701)20:10<2133::aid-elps2133>3.0.co;2-q
Subject(s) - dna , gel electrophoresis , electrophoresis , dna damage , comet assay , gel electrophoresis of nucleic acids , microbiology and biotechnology , chromatography , chemistry , biology , biochemistry
Single cell gel electrophoresis, also known as the comet assay, is widely used for the detection and measurement of DNA strand breaks. With the addition of a step in which DNA is incubated with specific endonucleases recognising damaged bases, these lesions can be measured, too. In the standard protocol, electrophoresis is carried out at high pH. If, instead, electrophoresis is in neutral buffer, the effect of DNA damage seems to be much reduced — either because alkaline conditions are needed to reveal certain lesions, or because the effect of the same number of breaks on DNA migration is greater at high pH. A lower sensitivity can be useful in some circumstances, as it extends the range of DNA damage levels over which the assay can be used. Here we compare the performance of standard and modified techniques with a variety of DNA‐damaging agents and offer possible explanations for the differences in behaviour of DNA under alternative electrophoretic conditions.