Dear Abdul Mannan,

Interesting question. Are you adding a 50mM stock solution to each well to end up with the concentrations you mention in your question? What is the volume you add of the 50mM stock solution to the wells with 100 microliter working volume, and are they all done with the same pipet? As I have calculated, you would have to add extremely small volumes to achive the concentrations you mention, which is often a source of variation which can not acoounted for. Kind regards, Harrie Verhoeven

Hi Harrie,

        Sorry for the confusion, 50mM is the stock in DMSO that I used to dilute it furrther.

I am diluting tamoxifen in the media for the required conc (double of final conc) and then adding 100 ul of it. 

Dear Abdul Mannan,

I suspect that during the dilution of the stock solution of 50mM in medium, at the highest concentration, the maximum solubility of Tamoxifen is locally reached, resulting in a partial precipitation (solubility in water  is 0.01%, ie 27 uM). If you then add the medium to the cells, it will not have the calculated concentration. I would like to suggest to sonicate the diluted stock in medium to dissolve the precipitate (5 min. or so), before adding it to the cells. I hope this helps. Kind regards, Harrie Verhoeven.

 Hi Harrie,

    Thanks you very much for your reply.

But, cell survival increases gradually with the increase in drug conc right from the 1uM conc. 

Attached file.

Thanks!