Large-Scale Production of High-Capacity Adenoviral Vectors

Begin with a storage bottle equipped with a magnetic stir bar containing a suspension of mammalian cells expressing Cre recombinase.
Co-infect the cells with a high-capacity adenoviral vector that lacks viral genes, allowing it to carry large foreign DNA inserts, along with a helper virus carrying the adenoviral genes.
Stir the suspension to promote viral and vector entry into the cells.
Transfer the cells to a high-volume capacity spinner flask containing pre-warmed medium and incubate under stirring to support large-scale vector production.
The helper virus encodes adenoviral proteins, while Cre recombinase removes its packaging signal to prevent helper virus encapsidation.
Meanwhile, the vector genome replicates and assembles into adenoviral particles using proteins supplied by the helper virus.
Transfer the infected cells to a tube and centrifuge to pellet the cells.
Discard the supernatant and resuspend the pellet in a small volume of buffer.
Store the infected cell suspension at a low temperature for downstream analysis.

Transfer the cells to a 250 milliliter storage bottle equipped with a sterile magnetic stir bar. Then, co-infect the cells with the passage three lysate and two trans