I am trying to culture S2R+ cells from frozen (liq. Nitrogen) aliquots. I am following the standard growth conditions as suggested on standard protocols, but have not been able to grow them properly. So, i would like to know if there are any enrichment that can be done for their optimum growth or do i just have to be patient and wait for them to grow ??
Thank you in advance for your valuable suggestions.
Is the thawed out culture very sparse? Do you have a lot of cells when you thaw them out? What are you thawing into, a T25? A lot of times cells just don't do well coming out of thaw, especially if they have been frozen for a while. You can increase the amount of FBS in your media and you can also thaw into a much smaller volume. Instead of a T25 you can thaw, wash and plate into 2 wells on a 6 well plate. Once those are confluent, pool those and and plate into 4 or 5 wells of the 6 well plate. If those grow well and are starting to look good, then you can expand them to a T25 or if you feel they are doing very well, a T75.
Hi Victoria, thanks for the suggestion. I plated three vials into one T25 and they were looking kind of semi-confluent. I would like to try the serum increament. So how much or what percentage do you think would be ideal to use?
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