Collins Andrew R
Department of Nutrition, University of Oslo, PB 1046, Blindern, 0316, Oslo, Norway.
Methods Mol Biol. 2017;1641:173-184. doi: 10.1007/978-1-4939-7172-5_9.
The comet assay is a sensitive electrophoretic method for measuring DNA breaks at the level of single cells, used widely in genotoxicity experiments, in biomonitoring, and in fundamental research. Its sensitivity and range of application are increased by the incorporation of an extra step, after lysis of agarose-embedded cells, in which the DNA is digested with lesion-specific endonucleases (DNA repair enzymes of bacterial or phage origin). Enzymes with specificity for oxidized purines, oxidized pyrimidines, alkylated bases, UV-induced cyclobutane pyrimidine dimers, and misincorporated uracil have been employed. The additional enzyme-sensitive sites, over and above the strand breaks detected in the standard comet assay, give a quantitative estimate of the number of specific lesions present in the cells.
彗星试验是一种用于在单细胞水平测量DNA断裂的灵敏电泳方法，广泛应用于遗传毒性实验、生物监测和基础研究。通过在琼脂糖包埋细胞裂解后加入额外步骤来提高其灵敏度和应用范围，该步骤中用损伤特异性核酸内切酶（细菌或噬菌体来源的DNA修复酶）消化DNA。已使用对氧化嘌呤、氧化嘧啶、烷基化碱基、紫外线诱导的环丁烷嘧啶二聚体和错配尿嘧啶具有特异性的酶。在标准彗星试验中检测到的链断裂之外，额外的酶敏感位点可对细胞中存在的特定损伤数量进行定量估计。
