CometAssay Assay Principle: R&D Systems

CometAssay Principal-How To Run Comet Assay

R&D Systems offers a complete set of reagents to run comet assay in our CometAssay^® and CometChip^® product lines. Our products include individual reagents, kits, electrophoresis units, and software. We provide everything you need to run a standardized comet assay and achieve reproducible results.

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What Is Comet Assay?

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. Comet assay is based upon the ability of denatured cleaved DNA fragments, or damaged DNA, to migrate out of the cell under electrophoresis creating a "comet tail", while the undamaged DNA remains within the cell membrane creating the "comet head." Comet assay is most commonly run under alkaline conditions to detect single- and double-stranded DNA breaks but can also be run under neutral conditions to detect only double-stranded DNA breaks.

Comet assay, single cell gel electrophoresis, is used to measure damaged DNA. Single lysed cells are run through electrophoresis.

How To Perform Comet Assay

Comet assay utilizes single cells to measure DNA damage. First, cells are embedded into agarose and then placed onto a slide. The slide is then immersed into lysis solution to break open the cell membrane. After the cells are lysed, DNA is denatured under neutral or alkaline conditions and run through electrophoresis. Cells are then stained with a fluorescent or chromogenic dye and visualized under a microscope. The resulting image obtained resembles a comet that includes a distinct head and tail. The comet head consists of undamaged DNA while the comet tail consists of the broken DNA strands. Analysis of the comet tail can be performed by hand or with software.

Untreated human Nalm6 cells were run through comet assay using CometChip.

Untreated human Nalm6 cells run on CometChip

Untreated human Nalm6 cells run on CometChip. Human Nalm6 cells were loaded into a CometChip and visualized using the CometChip Kit (Catalog # 4260-096-K) under alkaline conditions. No comet tail is observed due to samples being untreated.

Treated human Nalm6 cells run on CometChip where DNA damage is observed in the comet tail

Treated human Nalm6 cells run on CometChip. Human Nalm6 cells were first treated with 5 µM etoposide for 30 minutes and then loaded into a CometChip. Cells were visualized using the CometChip Kit (Catalog # 4260-096-K) under alkaline conditions. DNA damage is observed in the comet tail while undamaged DNA remains in the comet head.