Currently I am doing MTT assay with a well known anticancer drug Docetaxel and one compound of my interest for anticancer activity. I did experiment on MIa-PaCa2 and HeLA cell lines. I took 10000 cells per well and incubated them with various concentrations of drugs for 48 and 72 hours.
But I am facing a problem that positive control in my experiment, which is Docetaxel achieving IC50 in micromolar range but previous studies shown its IC50 in nanomolar range, I repeated the experiment many times but got the same results. I don't know why I am not achieving the IC50 in nanomolar range for Docetaxel. Can anyone help me in troubleshooting this problem?
Did the previous studies employ the MTT assay? Did you use the same MTT concentration and incubate for the same time?
Hi Elena, in one of the article, they have used WTS-1 based colorimetric assay. Should i use the same assay to achieve the IC50 value?
Rahul, yes, either use the WTS-1 colorimetric assay for a direct comparison (make sure all the conditions are the same as with the paper you are comparing) or find other studies which have used the MTT assay. This is one of the reasons you are not achieving the same IC50 concentration. Hope that helps.
I would consider the passage number of the cells and volume of the medium per each well.
You should consider the passage number,condition of cells, also check the media composition and if you are using antibiotics then also check their conc. All these factors also aafect the outcome results
Thank you all for your suggestions, i will keep these factors in mind next time.
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