Polybrene vs. Protamine Sulphate - (Jun/27/2007 )
I'm using polybrene currently as a transfection (and subsequently retroviral infection) agent in phoenix cells and HSCs respectively. The problem is polybrene is highly cytotoxic and i'm finding myself using more than the recommended amount (8ul in 1ml media per well of six well plate) in order to achieve sufficient transfection/infection efficiency, and as such i'm getting a lot of cell death. I've noticed in the literature that Protamine Sulfate is often used as an alternative (being a similar positively charged cation) but without the cytotoxicity, and i'm thinking about giving it a go. Does anybody have any experience in using protamine sulfate as an aid to retroviral infection?
Also, i've been using DMEM in my culture medium in which i've been infecting my cells and i'm concerned that the phenol red (which inhibits cation transporters in the cell) might be adversely effecting the efficiency of infection/transfection. Has anybody else found this to be a problem?
Any help is appreciated!
polybrene can aid in infection of retroviruses but this is applicable only for some cells. Anyway, It is toxic. If you have to use lots of polybrene which is killing cells, try increasing the amount of virus which you are adding.
I dont think phenol red in DMEM affects infection. We regularly infect cells in DMEM media without problems.
Good Luck !!!
We use polybrene as only in our virus stock. Once 293T are transfected and the virus ready, we collect the supernantant, filter it and add 4 ug/ml of polybrene. We do not add any more polybrene when infecting cells and we get a pretty good infection.