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Adaptation of mammalian cells to serum free mode


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#1 navya

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Posted 04 June 2011 - 09:41 PM

Hi

I am trying to adapt CHO stable transfected clones to serum free mobe.
While taking to low serum levels, ( from 10% FBS to 0.255 FBS) the doubling time has increased
I have followed sequnential adaptation method with 3 passages below 1% FBS level
Cells are not growinh in serum free mode in shaker.I have used Excell CD medium with feed supplements
How can i adapt them to serum free mode????


Navya

#2 klinmed

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Posted 06 June 2011 - 03:47 AM

Hi

I am trying to adapt CHO stable transfected clones to serum free mobe.
While taking to low serum levels, ( from 10% FBS to 0.255 FBS) the doubling time has increased
I have followed sequnential adaptation method with 3 passages below 1% FBS level
Cells are not growinh in serum free mode in shaker.I have used Excell CD medium with feed supplements
How can i adapt them to serum free mode????


Navya

Do your cells grow in SF medium when cultured in non-shake cultures (eg 25 cm2 flasks). If they do, you may need to turn the speed down. Cells in SF are very sensitive to shear forces. You could also try adding Pluronic F68 (final 0.1 %) to your medium).
Always remember that serum containing medium can "forgive many sins". With SF you must use very carefully cleaned glass vessels (preferably use disposable ones) and ALWAYS protect stock media bottles and cultures from light. If you still have problems you may need to use a slower adaption. For example, 5% serum to SF in 1% steps with 5 passages at each step.

Hope some of this helps

#3 navya

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Posted 25 June 2011 - 10:34 PM

Hey thanks so much ....
got my clones adapted to SFM mode!!!!!




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