Department of Biology, Faculty of Sciences, Autonomous University of Madrid, Spain.
Acta Histochem. 2012 Dec;114(8):785-96. doi: 10.1016/j.acthis.2012.01.006. Epub 2012 Feb 15.
Although MTT is widely used to assess cytotoxicity and cell viability, the precise localization of its reduced formazan product is still unclear. In the present study the localization of MTT formazan was studied by direct microscopic observation of living HeLa cells and by colocalization analysis with organelle-selective fluorescent probes. MTT formazan granules did not colocalize with mitochondria as revealed by rhodamine 123 labeling or autofluorescence. Likewise, no colocalization was observed between MTT formazan granules and lysosomes labeled by neutral red. Taking into account the lipophilic character and lipid solubility of MTT formazan, an evaluation of the MTT reaction was performed after treatment of cells with sunflower oil emulsions to induce a massive occurrence of lipid droplets. Under this condition, lipid droplets revealed a large amount of MTT formazan deposits. Kinetic studies on the viability of MTT-treated cells showed no harmful effects at short times. Quantitative structure-activity relations (QSAR) models were used to predict and explain the localization of both the MTT tetrazolium salt and its formazan product. These predictions were in agreement with experimental observations on the accumulation of MTT formazan product in lipid droplets.
虽然 MTT 被广泛用于评估细胞毒性和细胞活力，但 MTT 还原产物(formazan)的确切定位仍不清楚。在本研究中，通过直接观察活 HeLa 细胞和与细胞器选择性荧光探针的共定位分析来研究 MTT 甲臜(formazan)的定位。MTT 甲臜颗粒与线粒体没有共定位，这一点通过 rhodamine 123 标记或自发荧光显示出来。同样，MTT 甲臜颗粒与用中性红标记的溶酶体之间也没有观察到共定位。考虑到 MTT 甲臜的亲脂性和脂溶性，在用葵花籽油乳液处理细胞以诱导大量脂滴出现后，对 MTT 反应进行了评估。在这种情况下，脂滴中显示出大量的 MTT 甲臜沉积物。对 MTT 处理细胞活力的动力学研究表明，在短时间内没有有害影响。使用定量构效关系 (QSAR) 模型来预测和解释 MTT 四唑盐及其甲臜产物的定位。这些预测与 MTT 甲臜产物在脂滴中积累的实验观察结果一致。
