Tahara Haruna, Yamamoto Yusuke, Matsuda Shun, Yoshizawa Hiroe, Fujita Masaharu, Hioki Takanori, Kasahara Toshihiko
Safety Evaluation Center, Fujifilm Corporation.
J Toxicol Sci. 2017;42(2):205-221. doi: 10.2131/jts.42.205.
The ATP assay is a highly sensitive and versatile method for measuring cytotoxicity. However, the correlation between the cell viability results obtained using the ATP assay and those obtained using direct cell counting has not been widely reported. Therefore, to evaluate the reliability and limitations of the ATP assay, we compared the results of ATP assay with those of automatic cell counter, which can measure the number and diameter of cells directly, by using 24 compounds and repeating individual experiments thrice. The correlation between the data was low for 7 of the 24 compounds (r < 0.8, at least 2 out of 3 experiments). These were the top 7 of the 11 compounds that induced cell hypertrophy. These 7 compounds were also observed to increase the area of mitochondria. However, the last 4 of the 11 compounds increased the cell size but did not increase the mitochondrial area. For the remaining 13 compounds, which had no effect on cell size, a good correlation was observed between the results of the two methods (r > 0.8, at least 2 out of 3 experiments), and the cell size was effectively the same as that of the controls. We concluded that the poor correlation between the two methods was attributable to an increase in the content of intracellular ATP because of the chemically induced cell and mitochondrial hypertrophy. We showed that the ATP assay is unsuitable for assessing the cytotoxicity of compounds that induce cell hypertrophy with increase in the mitochondrial area and ATP content.
ATP 检测是一种用于测量细胞毒性的高灵敏度且通用的方法。然而，使用 ATP 检测获得的细胞活力结果与使用直接细胞计数获得的结果之间的相关性尚未得到广泛报道。因此，为了评估 ATP 检测的可靠性和局限性，我们通过使用 24 种化合物并将每个实验重复三次，将 ATP 检测的结果与自动细胞计数器的结果进行了比较，自动细胞计数器可以直接测量细胞的数量和直径。24 种化合物中有 7 种的数据相关性较低（r < 0.8，3 次实验中至少 2 次）。这些是诱导细胞肥大的 11 种化合物中的前 7 种。还观察到这 7 种化合物会增加线粒体的面积。然而，11 种化合物中的最后 4 种增加了细胞大小，但没有增加线粒体面积。对于其余对细胞大小无影响的 13 种化合物，两种方法的结果之间观察到良好的相关性（r > 0.8，3 次实验中至少 2 次），并且细胞大小实际上与对照相同。我们得出结论，两种方法之间的相关性较差是由于化学诱导的细胞和线粒体肥大导致细胞内 ATP 含量增加所致。我们表明，ATP 检测不适用于评估诱导细胞肥大并伴有线粒体面积和 ATP 含量增加的化合物的细胞毒性。
