I've checked two polybrene stocks (Sigma H9268-5G and one from Merck) in a titration of Bal virus on PBMCs and on TZMbls adding either polybrene to the medium or not. In both assays infection and titer goes down when I use polybrene. I used our in-house ELISA and the commercial Perkin Elmer one to measure p24 on PBMCs and I used Steadylite from PE for reading viral growth on TZMs. Anyway, this sounds a bit weird to me as the whole world is using polybrene to increase viral infection and in my ex-lab I did some experiments confirming this with Bal. Nevertheless my next step will be trying a different virus IIIB and SF162. Any reason why polybrene would affect different viruses differently? Anyone else having similar problems?
Hey Andrei
Thanks for your answer! Yeah, sorry, I should have been more complete/precise. I use 2µg/ml concentration as that hardly gives any toxicity on PBMCs in a polybrene titration exp w/o virus, on TZM there is a little bit toxicity as you can see this from the bit lower luciferase expression in the negative control but this is definitely not causing the observed effect. I did these experiments twice already on different days and different buffy coats. Haven't tried dextrane on PBMCs yet as that increases infection in TZMs....
What exactly do you mean with the logs?
Thanks
Phil
Hi
i think your cells make be undergoing stress on polybrene treatment. Decrease the concentration and change the media after some time, as polybrene help in entry so no need of it after that. I hope it will help you solve your problem.
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