Hello, I am going to make some in vitro incubation experiments (PBMCs in RPMI media) with estradiol (17-B), testosterone, and prednisolone.
I plan to dissolve these steroids in (200 proof) ethanol, and would like to prepare my stock in advance. Could I store them at -80C, or should I store them at -20? And do you know how long I could store this solutions under these temperatures?
I am buying these products from Sigma (E2758, T1500, P6004) , however, they are not endotoxin tested. Do you think I should use an 10kdalton filter to remove endotoxins? My worry is this filtration process could alter the final hormone concentration because these hormones are hydrophobic and might not %100 filter through the membrane? I see that a lot of people are using sigma products for their in vitro experiments, but I couldn't figure out if they are doing a protocol to remove the endotoxins.
Also, do anyone has any suggestion for storage veils for these hormones? I am looking for some glass veils, as I have read that Estradiol could be adsorbed by polypropylene.
4 C in ethanol or methanol in glass. No need to freeze. Won't freeze anyway. mark meniscus after using and top up ethanol if necessary
Steroids are quite stable - nevertheless we store them in a freezer (~ - 20°C), because evaporation of the alcohol is the critical issue (should be in tightly locked glass vials) - "losing" the alcohol would increase the concentration in the remaining stock solution.
Agree with Rupert. Fit of vials and lids is important. Don't store when diluted in aqueous solution.
Thank you very much Dr. Scott and Dr. Palme, these tips are very helpful!
Dear Osman Corbali, for cortisol we tested glass tube retention in Sadoul and Geffroy 2019, and I recommend that you Sodocalcic glass tubes: "The extraction is commonly performed in glass tubes, because steroids bind to plastic and because plastic is dis- solved by some solvents such as cyclohexane or ethylacetate. For this purpose, we recommend the use of sodocalcic rather than borosilicate glass, since we recovered on average 20% less cortisol from borosili- cate glass incubated with a cortisol solution (1 ng ml−1 in PBS–BSA). This difference between borosilicate and sodocalcic was significant (F = 5.8502, P < 0.05, n = 20) and is probably linked to the fact that sodocalcic glass is smoother due to a lower sand (SiO2) content."
Cheers
Ben
Sadoul, B. and Geffroy, B., 2019. Measuring cortisol, the major stress hormone in fishes. Journal of Fish Biology, 94(4), pp.540-555.
- Badges
- Science topic
- Similar topics
- Chemistry
- Pharmacology
- Pharmaceuticals
- Drugs