BPAE cells were plated in a 96 well plate, treated with 100 µM menadione for 1 hr ａnd then stained with 20 nM MitoTracker® Green, 5 µM Far Red ROS Sensor ａnd Hoechst 33342 for 30 mins in complete medium. Cells were then washed 3X with PBS ａnd imaged on a Thermo Fisher Cellomics ArrayScan® VTI. The control sample has no signal while in menadione treated cells, there is a robust increase in the signal as a result of oxidative stress caused by menadione. Both MitoTracker® Green ａnd Far Red ROS Sensor stained well showing that they work together in a multiplex assay.
Mitotracker正克服传统探针的缺点而被生产，其能够聚集在活线粒体上，并且细胞固定中也能良好维持。其中，Mitotracker orange，red，and deep red这三种探针经透化（permeabilization）后仍能良好维持。在接下来免疫细胞化学（ICC）、原位杂交（ISH）或电镜实验的处理步骤中样本仍保留线粒体的活细胞染色形态。另外，Mitotracker探针去除病原细胞线粒体染色可能遇到的一些问题，因为一旦线粒体染色后，细胞即可被固定然后进行结果分析。
【注1】：MitoTracker® Green FM（#M7514）在水溶液中无荧光； 【注2】：MitoTracker® Orange CM-H2TMRos（#M7511），MitoTracker® Red CM-H2XRos（#M7513）为还原态探针，无荧光。只有被氧化后才发荧光。
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