A comparative performance test of standard, medium- and high-throughput comet assays
A comparative performance test of standard, medium- and high-throughput comet assays
复制标题
DOI：
10.1016/j.tiv.2012.12.006
复制
发表时间：
2013-03-01
影响因子：
3.2
通讯作者：
Collins, Andrew R.
中科院分区：
文献类型：
--
作者：
Azqueta, Amaya;Gutzkow, Kristine B.;Collins, Andrew R.
传统彗星测定（单细胞凝胶电泳）的一个严重限制是电泳平台的大小限制了一次实验中可以处理的样品数量。提高通量的一种方法是减小凝胶的尺寸。我们在这里将显微镜载玻片上两个大凝胶的传统系统与最近的两项发展进行比较，即每个载玻片 12 个微型凝胶，以及 GelBond (R) 胶片上包含 96 个微型凝胶的格式。我们使用经过 X 射线或甲磺酸甲酯 (MMS) 处理的细胞。 X 辐射或 MMS 处理的细胞中检测到的损伤水平（% 尾部 DNA）不受所用形式的影响。使用所有三种形式的平行实验在两个独立实验室中使用 MMS 处理的细胞进行；两个实验室之间的结果差异具有临界意义。在凝胶边缘看到的异常彗星的潜在问题，即所谓的“边缘效应”，已经得到解决。可靠的高通量彗星测定可应用于基因毒性测试（特别是使用来自不同器官的样品进行体内研究）以及生态基因毒理学和人体生物监测，其中收集的样品数量可能相当大。 (C) 2013 Elsevier Ltd. 保留所有权利。
A serious limitation of the conventional comet assay (single cell gel electrophoresis) is the restriction on the number of samples that can be processed in one experiment, imposed by the size of the electrophoresis platform. One approach to increasing throughput is to reduce the size of gels. We here compare the conventional system of two large gels on a microscope slide, with two recent developments, namely 12 minigels per slide, and a format with 96 minigels on GelBond (R) film. We used cells treated with X-rays or methylmethanesulphonate (MMS). The level of damage detected (% tail DNA) in X-irradiated or MMS-treated cells was not affected by the format used. Parallel experiments, using all three formats, were performed with MMS-treated cells in two independent laboratories; the difference in results between the two laboratories was of borderline significance. The potential problem of anomalous comets seen at the border of the gel, the so-called 'edge effects', has been addressed. A reliable, high throughput comet assay has applications in genotoxicity testing (particularly for in vivo studies with samples from different organs) as well as ecogenotoxicology and human biomonitoring, where the numbers of samples collected can be considerable. (C) 2013 Elsevier Ltd. All rights reserved.