Comet assay, also known as single cell gel electrophoresis, is a sensitive technique for measuring DNA damage in single cells and has become well known in the field of toxicology and drug discovery. The Comet assay can detect specific DNA damage and repair, and thus can explore mechanism and mode of action to further investigate the effects of a compound.

 

The theory is that undamaged DNA retains a highly organized association with matrix proteins in the nucleus and when the DNA is damaged (by chemicals or UV radiation), this organization is disrupted. During the application of an electric field, the individual strands of DNA lose their compact structure and drift out of the nucleus and into the low melting point agarose suspension. The DNA (which has an overall negative charge) it is drawn towards the positively charged anode. The undamaged DNA does not move, whereas the damaged DNA (smaller fragments) are free to migrate away from the nucleus. Simply put, the amount of DNA which leaves the nucleus is a measure of the amount of DNA damage to the cell. The brighter and longer the DNA tail, the higher the level of damage.

The Comet Assay can be used to detect DNA damage caused by double strand breaks, single strand breaks, alkali labile sites, oxidative base damage, and DNA cross-linking with DNA or protein. The Comet Assay is also used to monitor DNA repair by living cells.