title: Adenovirus vector production using low-multiplicity infection of 293 cells.
abstract: The use of low-multiplicity infection of 293 cells in static culture with regular medium replacement was investigated for efficient large-scale production of adenovirus vectors for gene therapy applications. An adenovirus vector carrying the enhanced green fluorescent protein gene (Ad EGFP) was used to infect 293-F cells at a low multiplicity of infection (MOI) of 0.00001-0.1 transductional unit (TU) cell(-1). The cells, which have the ability to grow in suspension, were incubated in T-flasks and the serum-free culture medium was replaced with fresh medium via centrifugation every 2 days. Because only a small proportion of cells were initially infected at low MOIs (<1 TU cell(-1)), uninfected cells continued to grow until they were infected by progeny adenoviruses released from previously infected cells. When 293-F cells at a relatively low density of 1 x 10(5) cells cm(-3) were infected with Ad EGFP at a low MOI of 0.001 TU cell(-1), the vector yield was 2.7-fold higher than the maximum yield obtained with high-multiplicity infection (MOI = 10 TU cell(-1)) in batch culture. These results indicate that efficient adenovirus vector production using low MOIs is achieved by minimization of either nutrient depletion and/or accumulation of inhibitory metabolites in the culture medium.
authors: Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan.
authors: Kentaro
authors: K
authors: Yamada
authors: Naoya
authors: N
authors: Morishita
authors: Tomohisa
authors: T
authors: Katsuda
authors: Shuji
authors: S
authors: Kubo
authors: Akinobu
authors: A
authors: Gotoh
authors: Hideki
authors: H
authors: Yamaji