1 reply to this topic

[Rnotk]

Hi, All

I have few other questions about in vitro viral transfection.

so I do produce virus using producing cells and use the supernatant for transfection
and the question is regarding the serum content in medium during virus production and transfection.

someone told me before the presence of serum somehow interfere viral transfection, so since then I have used serum-reduced medium (1% serum instead of 10% without antibiotic) to collect virus,
and I also use same medium during transfection with the addition of polybrene.

but the problem is that I am using retrovirus, so with the reduced serum, proliferation of target cell slow down, which I think reducing tranfection efficiency for some of my experiment.

unfortunately my virus/expression vector does not have antibiotic resistance gene so I cannot do selection after the transfection.

So, my question is "Does serum really affect viral production/transfection???"

or if anyone know some way to increase trasnfection efficiency please let me know.

BTW, I tried polymer complex addition (polybrene/chondroitin sulfate), but my cell does not like it.

Thanks in advance

[protolder]

Rnotk, on 07 April 2011 - 08:52 AM, said:

Hi, All

I have few other questions about in vitro viral transfection.

so I do produce virus using producing cells and use the supernatant for transfection
and the question is regarding the serum content in medium during virus production and transfection.

someone told me before the presence of serum somehow interfere viral transfection, so since then I have used serum-reduced medium (1% serum instead of 10% without antibiotic) to collect virus,
and I also use same medium during transfection with the addition of polybrene.

but the problem is that I am using retrovirus, so with the reduced serum, proliferation of target cell slow down, which I think reducing tranfection efficiency for some of my experiment.

unfortunately my virus/expression vector does not have antibiotic resistance gene so I cannot do selection after the transfection.

So, my question is "Does serum really affect viral production/transfection???"

or if anyone know some way to increase trasnfection efficiency please let me know.

BTW, I tried polymer complex addition (polybrene/chondroitin sulfate), but my cell does not like it.

Thanks in advance

Hola, in Baculovirus medium without serum is indicated only in the trasfection phase with lipofectamine, and after 4 hours of entry of the complex into cells, you add serum and continue incubation untill the virus harvesting. but to me polietilenimine (JET Pei)is better; transfection could be done in serum media and the yield and price is better (at least for me). check in internet some of the references about your viruses and polietilenimine. Buena suerte