Talk:MTT assay

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Latest comment: 16 years ago1 comment1 person in discussion

like a very roundabout way to count cells! Is there some principled advantage to this assay over a direct count (with a haemocytometer (plus trypan blue), Coulter counter, flow cytometer, fluorescence microscope, etc), or is it a technical matter? This assay is presumably very easy to do on a 24-well plate and read with a plate reader; i wouldn't want to count twenty-odd samples on a haemocytometer.

The formation of the dye is catalysed by mitochondrial enzymes, so this assay really measures the quantity of those enzymes, right? That means you're not measuring cell number, but cell number times cell size times mitochondrial density times expression level of the enzymes times activity level of the enzymes. You have to assume that all the other factors stay the same. Do they? How would you know if they didn't? For example, aphidicolin blocks cell proliferation, but not growth, so cells treated with it will continue to get bigger, make more mitochondria, and develop a greater capacity to reduce MTT. If this assay tells you about the amount of mitochondrial activity, rather than the number of cells, is that a disadvantage, or is it actually an advantage, since it lets you detect things which make cells sick but don't stop them proliferating?

-- Tom Anderson 2008-02-01 2117 +0000

I agree with your comments. I have read a number of articles, some of which say that they used the MTT assay to measure cytotoxicity of different compounds, and others to measure cell growth and/or proliferation. It is possible that if cells are plated at confluency before the experiment, then any decrease in color change would be due to cell death and not a reduction in cell proliferation (assuming that mitochondrial density, enzyme expression and activity are constant - possibly not valid). However, most articles do not state what percentage confluency their cells are actually plated at prior to the assay, making it difficult to determine whether the results obtained were due to a change in proliferation rate (i.e. cell cycle length) or a change in cell death rates. In any case, I personally would not be happy plating cells at confluency unless I specifically wanted to monitor cell death of quiescent cells, because the propensity of cells to apoptose varies depending on their stage in the cell cycle.

I actually think that this assay is therefore quite misleading. Since we know that many drugs and pathways may impinge both on apoptotic and cell cycle processes, this assay does not allow you to delineate between these two effects. It should be used and interpreted with caution.

-- Adam Hardy 2008-09-16 [arhardy] —Preceding unsigned comment added by Arhardy (talk • contribs) 13:59, 16 September 2008 (UTC)Reply

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