I am working on bladder cancer cells. I am using Decitabine drug for my study. For experiments i dissolved drug in DMSO and after that i aliquoted it and stored it in -80. Every time i used a fresh aliquote for each experiment. I took all the precaution during experiment but i am not getting result even its effect is already reported in literature. could any one suggest me what changes i have to make for experiments
There could be so many reasons.
1. Try to use Decitabine from different vendors.
2. May be concentration you are using is too low for cytotoxic effects.
3. May be cell lines you are using is not sensitive to Decitabine.
4. You can also try other cytotoxic drugs like Paclitaxel which is very potent to make sure your experiment conditions are fine.
5. What is your experiment set-up? How are you measuring the efficacy of Dectabine?
See this paper from 2011 (http://www.landesbioscience.com/journals/epigenetics/Al-SalihiEPI6-8.pdf). Detection of effect is easy enough with western blotting even at early time points. You should find what you need in the materials & methods section. Decitabine is also water soluble, we prepared fresh before each experiment.
Please also note the effect of p53 status on decitabine action in the paper.
Dear Mazin
This is recommended in literature that we should use freshly prepared decitabine before each experiment. But constrain is that this drug is very costly and i have to add drug daily in culture medium if i am doing 72 hrs treatment in this case i can not afford making a fresh drug daily. So what i did was dissolve that drug in DMSO and stored it in -80 after aliquot it. And used new aliquot before each experiment. So is dissolving drug in DMSO and storing it in -80 , right procedure to use it for treatment point of view. So please suggest what can i do in this situation.
I would say the problem probably comes from your cells instead of decitabine. From by experience, cell culture contamination like mycoplasma has a huge effect on decitabine treatment.
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