Hello everyone..
Now, I work with ovine endometrial cell culture..
I will try to add progesterone to the culture medium. I currently have progesterone that can only be dissolved in absolute ethanol, while absolute ethanol will damage the medium. So, how is the right step to dissolve progesterone for ovine endometrial cell culture?
Thank you for your help,
Best regards, Ananda
Hi Ananda,
I suggest you first perform a test to see what percentage of ethanol is toxic to your cells (it will depend on the cells you are growing). Try a range betwen 0.1% and 5%. Then pick a reasonable dilution for your progesterone- so you can dilute your progesterone at very high concentrations so you only need to add a small amount of ethanol to your media. Of course for the control media without progesterone, still add ethanol as a control and you should be OK.
Dear Ananda,
try with N,N - DMF instead of ethanol.
Kind regards, Marko
Hi,
First you have to check the solubility of progesterone in different concentrations of ethanol then at the same time do toxicity test in your cells. Getting water soluble progesterone is also the best option for cell culture. good luck.
I agree with Ana Sowa that dilution in absolute ethanol would be the more suitable solution, provided that the final ethanol concentration after dilution in the medium remains lower than 0.1-0.5 %. Otherwise you may put a drop of pure ethanol-progesterone solution in the bottom of your culture flask, let it dry under sterile conditions and then add your medium and your cells. Complex between BSA and progesterone migt also be used, but the final concentration of progesterone in the medium and the kinetics of release can be difficult to appreciate.
You can make a stock P4 solution in mM range. This stock solution is stable for several months at -20 degrees. Than it is better to dilute the working (final) solution in PBS. The toxic effect of progesterone is negligible if the final concentrations of P4 in the culture medium are of the ranges of micro or nM.
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