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Lentivirus toxicity - (Oct/30/2010 )

Hi everyone,

Iīm trying to transduce mouse primary T lymphocytes with my empty vector (pLVTHM - GFP reporter). After the production of the viruses, I harvest the supernatant and centrifuge to remove cells/debris (2000rpm/10min) and filter it using the 0,22um syringe-driven unit from Millipore (PVDF membrane). However, after the ultracentrifugation step (using standard speed to concentrate VSV-pseudotyped vectors) I am obtaining a pellet (!), probably of cell debris or even proteins. Taking into account the size of the pellet, I hardly believe that it is formed only of viruses...If I ressuspend this pellet and use the virus to transduce the cells, two days later all my cells are dead. I didnīt discard the pellet because I was afraid to lose the viruses...

Any suggestions???

Leo Chicaybam


DO NOT use .22um filter, please use .45um filter before ultracentrifugation.

-Functional Screens-

Why not use the 0.22um filter?


Piersgb on Mon Dec 13 12:40:52 2010 said:

Why not use the 0.22um filter?

you will lose a lot of virus