Reliable Comet assay measurements for detecting DNA damage induced by ionising radiation and chemicals
Reliable Comet assay measurements for detecting DNA damage induced by ionising radiation and chemicals
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DOI：
10.1016/j.mrgentox.2006.03.002
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发表时间：
2006-06-16
影响因子：
1.9
通讯作者：
Jha, Awadhesh N.
中科院分区：
文献类型：
--
作者：
Kumaravel, T. S.;Jha, Awadhesh N.
单细胞凝胶电泳分析的碱性版本，通常称为彗星分析，广泛用于评估化学品和环境污染物的遗传毒性潜力，以及用于环境监测目的。近年来，该检测方法越来越被认为是一种潜在的有价值的监管研究工具。该测定通常利用市售软件程序来评估单细胞水平的DNA损伤程度。这些方案提供了大量的测量结果（即，尾长、%尾DNA、尾矩的各种测量等）以评估DNA迁移和DNA损伤的程度。然而，目前在最相关的测量或使用的参数方面没有达成普遍一致意见。本研究旨在确定彗星试验中哪些测量值与DNA损伤最显著相关，因此应用于常规使用。用一定剂量范围的（CS）-C-137 γ-辐射（0、1、2、4和8戈伊）辐射来自3个健康人个体的汇集的外周血样品。照射后，根据标准方案进行彗星测定，并通过使用Komet 5.0软件（Kinetic Imaging Ltd.，英国利物浦）。在相关性分析之后，Olive尾矩（OTM）、尾范围矩和尾中DNA的百分比（%尾DNA）给出了良好的相关性，其彼此没有显著差异。来自其他体外和体内化学试剂彗星试验的进一步回顾性分析也表明，OTM和%尾DNA与所用遗传毒性试剂的剂量具有良好的相关性。由于OTM和%Tail DNA是许多手稿中最常用的参数，因此这两个参数可以继续用于常规使用。然而，由于OTM是以任意单位测量的，并且不同的图像分析系统给出不同的值，因此可以认为%Tail DNA更有意义并且更容易概念化。在遗传毒理学研究中，其他参数可能不被认为具有重要意义。(c)2006 Elsevier B. V.保留所有权利。
The alkaline version of the single cell gel electrophoresis assay, popularly known as the Comet assay, is widely used to evaluate the genotoxic potential of chemicals and environmental contaminants, and for environmental monitoring purposes. In recent years, this assay has increasingly been recognized as a potentially valuable tool for regulatory studies. The assay commonly utilises commercially available software programmes to evaluate the extent of DNA damage at the single-cell level. These programmes provide a large number of measurement outcomes (i.e., tail length, %Tail DNA, various measures of tail moment, etc.) to evaluate the extent of DNA migration and DNA damage. At the moment, however, there is no general agreement with respect to the most relevant measurements or parameters to use. This study was carried out to establish which measurement(s) in the Comet assay are most significantly correlated with DNA damage, and should thus be adopted for routine use. Pooled peripheral blood samples from 3 healthy human individuals were irradiated with a range of doses Of (CS)-C-137 gamma-radiation (0, 1, 2, 4 and 8 Gy). Following irradiation, the Comet assay was performed according to a standard protocol, and different parameters were recorded by use of Komet 5.0 software (Kinetic Imaging Ltd., Liverpool, UK). Following a correlation analysis, the Olive Tail Moment (OTM), the Tail Extent Moment and the percentage of DNA in the tail (%Tail DNA) gave good correlations that were not significantly different from each other. Further retrospective analysis from other in vitro and in vivo Comet assay experiments with chemical agents also suggested that OTM and %Tail DNA gave good correlation with the dose of genotoxic agents used. Since OTM and %Tail DNA are the most commonly used parameters in many manuscripts, these two could continue to be applied for routine use. However, since OTM is measured in arbitrary units and different image-analysis systems give different values, the %Tail DNA could be considered more meaningful and easy to conceptualise. Other parameters might not be considered of significant use in genotoxicological studies. (c) 2006 Elsevier B.V. All rights reserved.