Limitations of MTT and MTS-based assays for measurement of antiproliferative activity of green tea polyphenols.
Limitations of MTT and MTS-based assays for measurement of antiproliferative activity of green tea polyphenols.
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DOI：
10.1371/journal.pone.0010202
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发表时间：
2010-04-16
期刊：
影响因子：
3.7
通讯作者：
Heber D
中科院分区：
文献类型：
--
作者：
Wang P;Henning SM;Heber D
绿茶多酚，如（-）-表没食子儿茶素-3-没食子酸酯（EGCG）的化学预防作用在细胞培养研究中已得到充分证实。然而，在不同实验室发表的关于EGCG抗增殖活性的研究中，观察到的半数抑制浓度（IC50）范围很广。尽管对EGCG处理的敏感性在很大程度上取决于癌细胞类型，但所采用的特定细胞活力和增殖检测方法可能会显著影响文献中报道的定量结果。
我们比较了五种广泛使用的方法，用于测量经EGCG处理后的LNCaP前列腺癌细胞和MCF - 7乳腺癌细胞的细胞增殖和活力。与台盼蓝检测相比，两种使用染料定量三磷酸腺苷（ATP）和脱氧核糖核酸（DNA）的方法在测量活细胞时都显示出准确性，并且结果显示出良好的线性相关性（r = 0.95）。然而，使用MTT（3 -（4,5 - 二甲基噻唑 - 2 - 基）- 2,5 - 二苯基四氮唑溴盐）和MTS（3 -（4,5 - 二甲基噻唑 - 2 - 基）- 5 -（3 - 羧基甲氧基苯基）- 2 -（4 - 磺基苯基）- 2H - 四氮唑）作为代谢活跃线粒体的指标，与ATP、DNA或台盼蓝测定相比，高估了活细胞的数量。结果，使用MTT和MTS测定的EGCG的观察到的IC50浓度比使用定量ATP和DNA的染料高2倍。相比之下，当细胞用芹菜素处理时，MTT和MTS检测与ATP、DNA或台盼蓝检测结果一致。
这些结果表明，基于MTT和MTS的检测方法会低估EGCG的抗增殖作用，并提示根据植物补充剂的化学性质仔细评估体外细胞活力和增殖评估方法的重要性。
The chemopreventive effect of green tea polyphenols, such as (-)-epigallocatechin-3-gallate (EGCG), has been well demonstrated in cell culture studies. However, a wide range of IC50 concentrations has been observed in published studies of the anti-proliferative activity of EGCG from different laboratories. Although the susceptibility to EGCG treatment is largely dependent on cancer cell type, the particular cell viability and proliferation assays utilized may significantly influence quantitative results reported in the literature. We compared five widely used methods to measure cell proliferation and viability after EGCG treatment using LNCaP prostate cancer cells and MCF-7 breast cancer cells. Both methods using dyes to quantify adenosine triphosphate (ATP) and deoxynucleic acid (DNA) showed accuracy in the measurement of viable cells when compared to trypan blue assay and results showed good linear correlation (r = 0.95). However, the use of MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) as indicators of metabolically active mitochondria overestimated the number of viable cells by comparison with the ATP, DNA, or trypan blue determinations. As a result, the observed IC50 concentration of EGCG was 2-fold higher using MTT and MTS compared to dyes quantifying ATP and DNA. In contrast, when cells were treated with apigenin MTT and MTS assays showed consistent results with ATP, DNA, or trypan blue assays. These results demonstrate that MTT and MTS -based assays will provide an underestimation of the anti-proliferative effect of EGCG, and suggest the importance of careful evaluation of the method for in vitro assessment of cell viability and proliferation depending on the chemical nature of botanical supplements.
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影响因子：
9.7
作者：
Thangapazham, Rajesh L.;Singh, Anoop K.;Maheshwari, Radha K.
通讯作者：
Maheshwari, Radha K.
影响因子：
--
作者：
O'Toole, SA;Sheppard, BL;Bonnar, J
通讯作者：
Bonnar, J
影响因子：
2.2
作者：
Jones, LJ;Gray, M;Singer, VL
通讯作者：
Singer, VL
DOI：
10.1124/jpet.103.049734
发表时间：
2003-07-01
影响因子：
3.5
作者：
Hsu, S;Bollag, WB;Schuster, G
通讯作者：
Schuster, G
影响因子：
--
作者：
Farabegoli, Fulvia;Barbi, Cristiana;Piva, Roberta
通讯作者：
Piva, Roberta