Many literature showed that how to determine IC50 drug concentration using MTT assay.
But I want to know how to determine non-cytotoxic concentration of a particular drug with maximum its biological activity using MTT assay i.e. Maximum concentration of drug with optimum biological activity but less cytotoxic effect.
Ideally, you should use flow cytometry and analyze apoptosis and cell cycle arrest at different drug concentrations aiming for that sweet spot (drug concentration) at which the cytotoxic effect is the lowest and your cytostatic effect is the highest (assuming that the biological activity of your drug somehow alters proliferation).
Using only MTT assay won´t let you discriminate between "biological" or cytotoxic effect, however, you could treat your cells with different concentrations of drug and identify the highest one at which there is no significant difference when compared to the vehicle (regarding MTT signal [metabolic activity]). Once identified, you could analyze the biological effect of your drug at that concentration using another approach (e.g. expression of genes regulated by the drug target, differences in signaling pathways, migration assays, invasion assays, etc.). If you see no difference between drug and vehicle groups you could try increasing the drug concentration slightly, considering that as you increase the drug concentration, the specificity of your test will decrease.
If this is known drug , then you can find the concentrations used for it on the Internet.
If this is a new drug, then make two or tenfold dilutions of this drug (depending on its activity) starting with the highest dose (for example, 10000, 1000, 100, 10, 1, 0.1) and perform an MTT test with control (cells without drug). Based on the results of MTT, build a graph of the dependence of the dose of the drug - cell death, you will determine the lowest toxic dose of this drug.
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