I am trying to express a protein in HEK293 freestyle. But I have some dilemmas regarding culturing HEK 293F in spinner flask. Although cells are dividing rapidly in tissue flask (T75 and T225), they start to die having cultured them in spinner flask.
Temperature is 37
CO2 level: I tried both 5% and 8%
stirring speed: 100 - 110 rpm
Cell density for seeding 200 k - 400 k cell/mL
Media: HEK FreeStyle Expression Media
I think there is no issue with incubator temperature or CO2 level, since as I said earlier cell grow normally in a T225 flask using the same incubator. I think the problem might emanate from spinner flask and speed. I was wondering to ask someone who has experience using HEK293 freestyle (or similar cells) and had the same issue?
Dear Shahryar
to growth HEK293 in suspension i suggest to you to use cap vented shake sterile shake flask.
For example we using the 125ml ( VWR code 734-1885) for growth 30ml or the 250ml to growth 60ml of cells.
We are growing Expi293 (which are similar to HEk293) in this flask and in shake incubator at 37°C, 120rpm, CO2%.
good luck
Manuele
Dear Shahryar
I am working with Expi293 cells for a while. I grow them in 125 ml cap vented sterile shake flasks as recommended and I never tried using tissue flasks. My culture volume is 30mL for 125ml flask. I prefer seeding density between 300-400k/mL and I never let them grow more than 3x10^6/mL. Our incubator condition is 37C, 125rpm and 8% CO2.
Good luck
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