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Will pervious PBS concentration in medium have a long lasting effect on cell gro


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#1 hello world

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Posted 23 February 2011 - 02:57 AM

Hi all,

(the topic is wrong, it should be FBS not PBS!~)

I have transfected NIH 3t3 with gene of interested and selected with G418. The process has repeated twice. For the first time of transfection, most of the cells die due to low confluency during selection. In order to rescue the transfected HEK, i'vd rised 10%FBS (+90% DMEM) to 20% FBS for 2 weeks in absence of G418. When it grows back to normal 60% confluency, i use back 10% PBS and select again using G418 for at least 2 week. (for the second time of transfection, the process is smooth and no change in % of FBS)

For both transfected 3t3, gene expression is checked and are found to be similar among these 2 transfected cell line. I have done the proliferation assay for these 2 indepedently transfected 3t3(toegther with postive and negative control).

The problem is that the one cultured with 20% PBS perviously have similar result with postive control while another transfected cell get result close to negative control in the proliferation assay (MTS).

I dont know whether such difference of transfected cells with similar expression of gene of interested is caused by long lasting effect of 20% PBS or viral contamination (for the transfected cell with result closed to negative control, the medium SEEMS to be more susceptibleto turn yellow, but the 3t3 morphology like totally fine and grow well).

Therefore, can anyone tell me whether pervious PBS concentration in culture medium will really have a long lasting effect on cell growth?


Cheers,
hello world

Edited by hello world, 23 February 2011 - 08:03 AM.


#2 tonghuitong

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Posted 27 February 2011 - 04:39 AM

I think you need not change the concentration of FBS, just keep it 10%.
otherwise, your results can not be convinced.

#3 hello world

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Posted 02 March 2011 - 07:29 AM

ok~ thank you for your reply.




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