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  • Title: Baculovirus production for gene therapy: the role of cell density, multiplicity of infection and medium exchange.
    Author: Carinhas N, Bernal V, Yokomizo AY, Carrondo MJ, Oliveira R, Alves PM.
    Journal: Appl Microbiol Biotechnol; 2009 Jan; 81(6):1041-9. PubMed ID: 18923829.
    Abstract:
    One of the major concerns regarding the use of insect cells and baculovirus expression vectors for the production of recombinant proteins is the drop in production observed when infecting cultures at high cell densities; this work attempts to understand this so-called cell density effect in the scope of baculovirus production for gene therapy purposes. A Spodoptera frugiperda insect cell line (Sf-9) was cultured and infected in serum-free medium, and the patterns of production of a recombinant baculovirus expressing the green fluorescent protein (GFP) were analyzed at different cell concentrations at infection (CCIs) and multiplicities of infection (MOIs). The results confirm that a cell density effect on productivity occurs which is dependent on the MOI used, with a high MOI "delaying" the drop in production to higher cell densities. Medium replacement at the time of infection using a high MOI considerably improved baculovirus production, with the different production indicators, namely the titer, specific yield, amplification factor, and time of harvesting, increasing with cell concentration for the CCI range tested. Virus titers as high as 2.6 x 10(10) IP x mL(-1) were obtained in cultures infected at 3.5 x 10(6) cells x mL(-1), while the amplification factor was roughly 19 times higher than the highest value obtained without medium exchange.
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