A Versatile Cell Death Screening Assay Using Dye-Stained Cells and Multivariate Image Analysis
A Versatile Cell Death Screening Assay Using Dye-Stained Cells and Multivariate Image Analysis
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DOI：
10.1089/adt.2015.661
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发表时间：
2015-11-01
影响因子：
1.8
通讯作者：
Andrews, David W.
中科院分区：
文献类型：
--
作者：
Collins, Tony J.;Ylanko, Jarkko;Andrews, David W.
提出了一种新的基于染料的方法，用于在基于图像的屏幕中测量细胞死亡。与仅准确测量一种形式的细胞死亡的传统高通量和中通量细胞死亡测定不同，使用对用廉价混合物、红色染料壬基吖啶橙和核染色剂染色的细胞显微照片进行多变量分析，即使没有阳性对照，也可以量化由各种不同激动剂诱导的细胞死亡。令人惊讶的是，使用单一已知的细胞毒性剂作为训练多变量分类器的阳性对照，可以准确量化机械上不相关的化合物的细胞毒性，从而能够生成剂量反应曲线。与低通量生化方法的比较表明，细胞死亡与低浓度生物活性化合物衣霉素和布雷菲德菌素 A 诱导的细胞应激能够准确区分。对 300 多种激酶抑制剂进行高通量基于图像的格式分析，正确识别出 11 种激酶抑制剂具有细胞毒性，只有 1 种假阳性。这种基于染料的测定的简单性和稳健性使其特别适合活细胞筛选有毒化合物。
A novel dye-based method for measuring cell death in image-based screens is presented. Unlike conventional high- and medium-throughput cell death assays that measure only one form of cell death accurately, using multivariate analysis of micrographs of cells stained with the inexpensive mix, red dye nonyl acridine orange, and a nuclear stain, it was possible to quantify cell death induced by a variety of different agonists even without a positive control. Surprisingly, using a single known cytotoxic agent as a positive control for training a multivariate classifier allowed accurate quantification of cytotoxicity for mechanistically unrelated compounds enabling generation of dose-response curves. Comparison with low throughput biochemical methods suggested that cell death was accurately distinguished from cell stress induced by low concentrations of the bioactive compounds Tunicamycin and Brefeldin A. High-throughput image-based format analyses of more than 300 kinase inhibitors correctly identified 11 as cytotoxic with only 1 false positive. The simplicity and robustness of this dye-based assay makes it particularly suited to live cell screening for toxic compounds.