Reading many papers I´ve found that in this context (cell proliferation) using metabolic assays such as MTT is like "the gold standard". Many papers use this approach when trying to determine inhibition of proliferation of an anticancer drug, but i have also noticed that many persons from the scientific community say that this is not the best approach and that a DNA dye-based assay will be more "representative" of cell proliferation inhibition.
I´ll like to know which is your opinion in this regard and what you will recommend using and why.
I´m working with antiandrogens in cancer cell lines.
MTT assay is based on the activities of mitochondrial dehydrogenases and is used as an indirect method of cellular proliferation. On the other hand DNA replication based techniques such as BrDU assay is also used as an alternative approach. Another easy and cheap way is Trypan Blue Assay.
Sometimes it is possible that the cell has halted the DNA replication due to DNA damage, as many chemotherapeutic agents causes cell cycle arrest, the cell will resume the cycle is dependent on the severity of DNA. If it's reparable than the DNA replication and cell cycle may resume. Thus it is advisable to choose the method carefully.
As MTT, MTS and XTT all uses mitochondrial dehydrogenase to cleave a substrate to produce a metabolic product to be measured, some drugs that affect metabolism (e.g. drugs that inhibit NAD+ dependent enzymes) may make the assay inaccurate. If this is the case, the use of other DNA binding dyes (for example CyQuant) may improve the assay credibility.
Anticancer drugs exert their cytototoxic effects by different ways. MTT assay as mentioned by others actually measures the metabolic activity of cells based on the mitochondrial dehydrogenase activity. Deregulated metabolic activity might affect the cell cycle progression. However, certain class of drugs can directly affect the cell cycle such as Doxirubicin, paclitaxel etc. In these cases determination of cell cycle progression is more logical. Again cell cycle arrest might cause activation of mitochondrial apoptosis and thereby reduced mitochondrial activity. Therefore, depending on the time of analysis, both the classes of drugs can cause changes in metabolic activity as well as in cell cycle. However, if you think that your drug is primarily affecting DNA replication or cell division then go for Cell cycle analysis otherwise MTT.
Cheers
As before mentioned others colleagues MTT, MTS and XTT all uses mitochondrial dehydrogenase and there are used as an INDIRECT measure of proliferation, I Think it is cellular viability. If you want measure DNA content, that is DIRECTLY related to cellular proliferation, you must use DNA binding dyes.
Good luck.
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