When making lentivirus, after harvesting the virus, do you usually filter them with 0.22um or 0.45um filters? Most protocols use 0.45um, but I was wondering would 0.22um hurt? What is the size of the lentiviral particles? It says 80-100nm on wikipedia, which means that the 0.22um filter is large enough and should not decrease the titer of the virus? Thanks for the help!
Dear Stephan,
Absolutely 0.45µm, for two reasons: 1) 0.22µm or 0.45µm is the maximum pore size, most of the pores are smaller; 2) there are so many cell debris which may block the pores, resulting for loss of lentivirus.
Genemedi got a rich experience in lentivirus production with Lipo transfection reagent, you could find more information on https://www.genemedi.net/i/lentivirus-packaging
Hope this may help you.
As you noted, the lentivirus ranges between 80–100 nm in diameter. Therefore, both types of filters can be used. However, it is common to use 0.45 micrometer (or 450 nm) size filter. The other filter could get clogged easily due to the lower size limit.
However, Cellecta (a company specialized in producing lentiviruses) recommends filtering through 0.22 PES membranes. https://www.cellecta.com/wp-content/uploads/Cellecta-Manual-CRISPR-Pooled-sgRNA-Libraries.pdf. I don't think 0.22 PES filters (PES is very low binding membrane) will significantly decrease virus titers. Once I get any data I will come back to feedback.
Hi Angel L. Alvarez , I'm interested to know how you went with the 0.22 PES filters?
Cindy Hsin-Yi Tseng I am having very low titers with both 0.45 and 0.22. The titers do not differ greatly among them though. At the end, I had to concentrate anyway to increase titers regardles the filter I used. That been said, I think 0.45 um is more than enough to filter lenti preps for what it is: removing cell debris. Hope this could help you.
Angel L. Alvarez Good to know! We ran out of 0.45um filter before and luckily we managed to borrow some from another group. At least 0.22um may be an option if this happened again.
Cindy Hsin-Yi Tseng By the way, regardles the pore size , use only cellulose acetate or polyethersulfone (PES) (low protein binding) filters. Do not use nitrocellulose filters as nitrocellulose binds surface proteins on the lentiviral envelope destroying the virus.
Dear Stephan,
Absolutely 0.45µm, for two reasons: 1) 0.22µm or 0.45µm is the maximum pore size, most of the pores are smaller; 2) there are so many cell debris which may block the pores, resulting for loss of lentivirus.
Genemedi got a rich experience in lentivirus production with Lipo transfection reagent, you could find more information on https://www.genemedi.net/i/lentivirus-packaging
Hope this may help you.
In fact, a 0.45 filter has a filtration rate of 75%, and a 0.22 filter has a 60% filtration rate.
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