I have cultured iPSC on MEF feeder layers. 2-mercaptoethanol is one of components of iPSC media. When I make iPSC media, I put 0.1 millimolar 2-mercaptoethanol into media and use it during experiment(maybe one month). However, I saw one of answer that half life of 2-mercaptoethanol is very short(I heard in 1 day) even though it is stored at 4℃. If it is true, how do you manage to keep concentration of 2-mercaptoethanol? I worry about a situation that effect of 2-mercaptoethanol is gone because of short half life. Thank you for reading my question.
We used to use BME in our stem cell/iPSC media. It was taken out since people did not add it fresh daily. It makes no difference in the culture conditions. BME is one of those things people add just because others in the past did. It is not necessary if you are using defined media.
Fresh addition of 2-mercaptoethanol to your iPSC media would help!!
It is very simple, one needs to add 2-mercaptoethanol fresh into the medium each time you change the medium (say every other day). If not, then add based on its know half-life at 37 degree C.
Dear. Gangaraju and professor, Choubey.
I approciate your advice. I will do it. I have 14.3M of 2-mercaptoethano. After obtaining your advice, I think I will make its stock and use it. Thank you again.
I would also suggest supplying your media with fresh ß-mercaptoethanol every day, as you need to do daily media changes for iPSC either way :) Good luck.
Thanks, Günes. I will do it according to your advice. That's best way to keep good conditions for iPSC.
2-ME is an anti-oxidant and you're right. The half life is very short. You need to protect your medium from light. In my ESC or iPSC cultures, I add them whenever I prepare medium (35 µL/50 mL PBS and 1%). I prepare medium typically 2-3 times a week. You cannot keep 2-ME stock and should prepare fresh (35 µL/50 mL) every time. Hope this is helpful.
Dear. Jung
Thank you your advice. I have a question and a concern. One of your advise is not to keep 2-ME stock. Because of short half life? When I make a medium, I use it for near 25days since my samples are a few. After obtains many advices, I think I aliquot medium without 2-ME and then put 1mM of 2-ME into it for working concerntraion. Is it bad idea?
Yes and no. I prepare stem cell culture media only 100-200 mL and use them in the following 3-4 days.
Dear. Jung
Thank you. I refer your advice.I should change my experiment schedule.
We used to use BME in our stem cell/iPSC media. It was taken out since people did not add it fresh daily. It makes no difference in the culture conditions. BME is one of those things people add just because others in the past did. It is not necessary if you are using defined media.
Dear. Kulik
Thank you for giving your advice. I don't worry about that BME works or not after your answer. Thank you again.
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