HELIX3
INC.
The Comet Assay
Description
The comet or single cell gel (SCG) assay measures DNA damage (e.g. strand breaks, DNA adducts, excision repair sites, and crosslinks) at the single cell level. Without the necessity for special cell isolation techniques, a small amount of cells is embedded in an agarose gel matrix on a microscope slide and lysed to remove the cell membrane and most of the cellular proteins.
Important Information
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Types of DNA Damage Detected
DNA migration is measured by image analysis
Cells are embedded in agarose on a slide
Lysis removes cell membranes leaving only nuclear DNA on slides
Alkaline (pH>13) buffer denatures DNA to express strand breaks
DNA pieces are migrated away from the nucleus
Strand Breaks
DNA Adducts
Crosslinks
Oxidative Damage
Once the cells are lysed, an electric current is applied to the nuclear DNA causing DNA fragements to migrate away from the nucleus or the head of the comet at a rate and distance directly proportional to the size of the fragment. The pixel intensity of the migrated fragments can then be measured to determine the DNA migration levels as an indication of damage and/or repair in a population of cells.
