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MitoTracker™ Deep Red FM - Special Packaging, 20 x 50 μg, 20 x 50 μg - FAQs

MitoTracker™ Deep Red FM - Special Packaging, 20 x 50 μg, 20 x 50 μg - FAQs

查看更多产品信息 MitoTracker™ Dyes for Mitochondria Labeling - FAQs (M22426, M22425, M7514, M7512, M7513, M7510, M7511)

7 个常见问题解答

我用MitoTracker Red CMXRos标记神经元后，它们第二天就死了，这是否正常？

Mitotracker染料应在染色之后立即成像，因为随着时间的推移，这些染料可能会产生毒性。

当我测试线粒体膜电位时，未处理的细胞也会发出荧光，而且我在试验样品中没有看到显著差异。

无论您使用哪种染料，四甲基罗丹明甲酯（TMRM）还是MitoTracker Red FM，未处理的细胞都会发出荧光。只要细胞线粒体膜电位降低就会导致荧光信号降低。最重要的是变化的程度。JC-1染料不仅强度改变，还有激发和发射比例光谱的改变。设置未处理的对照和用线粒体膜电位去稳定剂（如CCCP或FCCP）处理的阳性对照是非常重要的。这些染料仅用于活细胞，在固定处理的细胞中无法保留相同程度的信号。

It looks like my Mitotracker dye is staining more than just the mitochondria. Why?

This is typically a result of using too high of a concentration of the Mitotracker dye. Most organic dyes are used in the low micromolar range. The MitoTracker dyes are used at a much lower concentration, around 50–200 nanomolar. Higher concentrations can cause background fluorescence and non-mitochondrial staining.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Can I use MitoTracker Red CMXRos dye in a plate reader to study mitochondrial membrane potential changes?

Yes, it has been done. A literature search will find numerous examples. However, be aware that plate readers are typically less sensitive than microscopes or flow cytometers, which means that the degree of change in fluorescence may not be as sensitive or easily detectable with a plate reader. Be sure to optimize label times and concentrations carefully.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Our facility does not allow flow cytometry of live cells. Can I fix the cells after staining with MitoTracker Deep Red FM dye?

Yes, this is possible because MitoTracker Deep Red FM dye is retained upon fixation. See Table 2 in this manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/mp07510.pdf).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

After I labeled neurons with MitoTracker Red CMXRos, they are dead the next day. Is this expected?

The Mitotracker dyes should be imaged soon after staining because over time, those dyes can be toxic.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I am testing mitochondrial membrane potential, but my untreated cells are fluorescing, and I'm not seeing a significant difference in my test sample.

Regardless of which dye you use - tetramethylrhodamine, methyl ester (TMRM), JC-1 or MitoTracker - untreated cells will fluoresce. It's just that cells with reduced mitochondrial membrane potential will fluoresce less. It is the degree of change which is important. JC-1 dye not only changes intensity, but has a ratiometric spectral change in excitation and emission. It is very important to have an untreated control as well as a positive control treated with a mitochondrial membrane potential destabilizer, such as CCCP or FCCP. Most mitochondrial stains are only for use with live cells, as the signal will not be retained to the same degree with fixation.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.