The Alkaline Comet Assay, also known as the Single Cell Gel Electrophoresis (SCGE) assay, is a widely used laboratory technique employed in the field of genetic toxicology. It is used to evaluate DNA damage and repair in individual cells.
In this assay, cells are embedded in a gel matrix and then lysed to release their DNA. The DNA molecules, being negatively charged, migrate towards the anode when an electric field is applied. However, damaged DNA moves slower than intact DNA due to breakages or lesions in the DNA strands.
The distinction between damaged and undamaged DNA is visually observed by staining the DNA with a fluorescent dye. The resulting migration patterns appear like comet tails, with intact DNA forming the 'head' and damaged DNA forming the 'tail'. These comet formations quantitatively indicate the level of DNA damage, with longer tail lengths indicating greater damage.
The assay can be tailored to assess different types of DNA damage by altering the conditions, such as using enzymes to target specific types of DNA lesions. Additionally, by incorporating measures to inhibit DNA repair mechanisms, the assay can also be used to evaluate DNA repair efficiency.
The Alkaline Comet Assay finds applications in various fields, including environmental monitoring, molecular epidemiology, pharmaceutical and cosmetic industry research, and genotoxicity testing of chemical compounds. It serves as an essential tool for assessing the genotoxic potential of substances and understanding the mechanisms behind DNA damage and repair.
