Continuous harvesting of lytic viruses using a VHU perfusion system

Lytic viruses such as Influenza A Virus are normally produced using adherent cell processes since suspension cell processes display a “cell density” effect where the cell-specific virus yield (virions/cell) decreases significantly as the cell density increases beyond 1-5 million cells/mL.  Prior investigations have shown that the cell-specific virus yield can be sustained with increasing cell density using a perfusion system consisting of a virus harvest unit (VHU®, Cambridge, MA) which allows for continuous nutrient supply and removal of toxic compounds. Virus titers more than 10 times higher in perfusion mode compared to previously reported batch cultivations were observed as a result of achieving similar or higher cell-specific virus yield (~50-90 PFU/cell) at cell densities increasing from 2×106 cells/mL in batch to 2×107 cells/mL in perfusion. During the virus replication phase, the VHU®, in contrast to the hollow fiber membrane, allowed 100% of the produced virus particles to pass through into the permeate including cell debris and host cell proteins as well as host cell DNA released with cell lysis. The VHU® appeared not to retain any protein.  In strong contrast, a hollow fiber membrane filter was shown to withhold most proteins which may result in greater fouling of the membrane over time. Overall, the VHU® perfusion process offers interesting possibilities for advanced process integration strategies for next-generation virus manufacturing.