How does alkaline comet assay work?
The alkaline comet assay measures DNA strand breaks at the single-cell level. Suspensions of single cells are embedded in agarose on a microscope slide and the cells lysed to form nucleoids, which contain supercoiled loops of DNA.
What does alkaline comet assay measure?
The alkaline comet assay (single cell gel electrophoresis) is the most widely used method for measuring DNA damage in eukaryotic cells (Neri et al., 2015).
Why alkaline comet assay?
The alkaline comet assay is capable of detecting DNA double-strand breaks, single-strand breaks, alkali-labile sites, DNA-DNA/DNA-protein cross-linking, and incomplete excision repair sites.
How do you do a 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.
What does the comet assay detect?
Comet assay is a microgel electrophoresis technique, which detects DNA damage and repair in individual cells. The assay measures DNA damage (i.e., strand breaks, DNA adducts, excision repair sites, and cross-links) at the single-cell level.
What is the purpose of comet assay?
The comet assay is a versatile technique for detecting damage and with adjustments to the protocol can be used to quantify the presence of a wide variety of DNA altering lesions (damage). The damage usually detected are single strand breaks and double strand breaks.
What does comet assay detect?
What does comet assay stand for?
single-cell gel electrophoresis
The comet assay (single-cell gel electrophoresis) is a simple method for measuring deoxyribonucleic acid (DNA) strand breaks in eukaryotic cells. Cells embedded in agarose on a microscope slide are lysed with detergent and high salt to form nucleoids containing supercoiled loops of DNA linked to the nuclear matrix.
Why comet assay is called single cell gel electrophoresis?
The Comet Assay, also called single cell gel electrophoresis (SCGE), is a sensitive and rapid technique for quantifying and analyzing DNA damage in individual cells. Individual cells are embedded in a thin agarose gel on a microscope slide. All cellular proteins are then removed from the cells by lysing.
Is there a validation study for the in vivo comet assay?
The JaCVAM international validation study on the in vivo comet assay: Selection of test chemicals The Japanese Center for the Validation of Alternative Methods (JaCVAM) sponsored an international prevalidation and validation study of the in vivo rat alkaline pH comet assay.
Are there any carcinogens in the jacvam test?
These 40 chemicals included 19 genotoxic carcinogens, 6 genotoxic non-carcinogens, 7 non-genotoxic carcinogens and 8 non-genotoxic non-carcinogens. “Genotoxicity” was defined as positive in the Ames mutagenicity test or in one of the standard in vivo genotoxicity tests (primarily the erythrocyte micronucleus assay).
What was the jacvam international validation study on in vivo?
The Japanese Center for the Validation of Alternative Methods (JaCVAM) sponsored an international prevalidation and validation study of the in vivo rat alkaline pH comet assay.