Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of radiosensitivity.
Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of radiosensitivity.
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发表时间：
1987-02
期刊：
影响因子：
11.2
通讯作者：
J. Carmichael;W. Degraff;A. Gazdar;J. Minna;James B. Mitchell
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作者：
J. Carmichael;W. Degraff;A. Gazdar;J. Minna;James B. Mitchell
使用标准克隆形成试验、染料排除试验和利用四唑盐、3-（4，5-二甲基噻唑-2-基）-2，5-二苯基溴化甲瓒的半自动比色试验，分别生成倍增时间为10、20和85 h的V79中国仓鼠和两种人肺癌细胞系（NCI-H460和NCI-H249）的辐射存活曲线。在倍增时间为10和20 h的细胞系中，观察到所有试验的D 0和外推数（n）结果相当。在这些情况下，肿瘤细胞系在辐射后经历了七次或更多次的倍增。对于倍增时间为80 h的肿瘤细胞系（H249），两种试验之间的D 0 S相当，而3-（4，5-二甲基噻唑-2-基）-2，5-二苯基溴化甲瓒试验中的外推数增加，结果可能与辐射后倍增数较低（小于4）有关。然后，我们使用已知试剂测试了测定检测辐射防护和致敏的能力。我们发现，半胱胺处理导致辐射防护（在8戈伊时为8倍），而5-溴-2-脱氧尿苷掺入导致所有三种测定中的辐射敏感性增强。我们的结论是，虽然必须建立每个细胞系的最佳条件（细胞数平板和倍增时间），使用特征的肿瘤细胞系，3-（4，5-二甲基噻唑-2-基）-2，5-二苯基溴化甲瓒测定可以自动化，因此在筛选大量潜在的放射增敏剂或保护剂中具有很大的价值。
Radiation survival curves were generated for V79 Chinese hamster and two human lung cancer cell lines (NCI-H460 and NCI-H249) with doubling times of 10, 20, and 85 h, respectively, using a standard clonogenic assay, a dye exclusion assay, and a semiautomated colorimetric assay utilizing a tetrazolium salt, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylformazan bromide. Comparable results for D0 and extrapolation number (n) were observed for all assays in the lines with doubling times of 10 and 20 h. In these instances the tumor cell lines had undergone seven or more doublings after radiation. For the tumor line (H249) with an 80-h doubling time the D0S were comparable between the assays while the extrapolation number was increased in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylformazan bromide assay, a result probably related to the lower number of doublings (less than 4) after radiation. We then tested the ability of the assays to detect radiation protection and sensitization using known agents. We found that cysteamine treatment resulted in radioprotection (by a factor of 8 at 8 Gy) while 5-bromo-2-deoxyuridine incorporation caused enhancement of radiation sensitivity in all three assays. We conclude that, while optimal conditions for each cell line (cell number plated and doubling time) must be established, using characterized tumor cell lines, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylformazan bromide assay could be automated and thus be of great value in screening large numbers of potential radiosensitizers or protectors.