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sf9 Insect cells questions


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3 replies to this topic

#1 saad

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Posted 26 May 2012 - 05:58 AM

Hi all,

I'm working on the sf9 insect cells to generate recombinant baculovirus for protein expression.
I use bac to bac system. The cells are growing (bit slower) for now about 4-5 weeks, I've noticed some balck dots in the media and granulles inside the cells. Is it sign for conatmination? I'm still diluting the cells and the doubling time is around 48 hours and sometimes less and some times a bit longer. The cells seem to pass passage 30.
If the viability of cells are high and still growing, what are the sign for contamination in my case.
I did the transfection and the WB showed some expression so I want to ampilfy the virus. I'm wondering about the cells if they are healthy and good for propagating the virus and protein expression later. The vaiability is > 95% with trypan blue exclusion.
Any suggestion or comments are highly appreciated.
Thank you in advance for your help as usual.

#2 hln

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Posted 27 May 2012 - 12:57 AM

contamination. start over.

#3 saad

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Posted 27 May 2012 - 02:27 AM

Thank you hln
Do you mean that the small balck dots are some type of bacteria. I'm using the same hood that been used by other people for mammalian culture.
I've heard about mycoplasma. Also, If the cells are contaminated do they still growing in that case.

Thank you so much,

#4 hln

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Posted 27 May 2012 - 11:13 PM

Yes, they're likely bacteria. Your cells may or may not continue to grow. In any case, you don't want to continue working with them. You will get suboptimal, non-reproducible results. If other people are also getting contamination, you need to clean the hood.




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