4.2 Multiplicity of Infection (MOI) of Baculovirus Vectors
In the IC-BEVS, the MOI of the baculovirus has a critical impact on the rate of cell growth and viability, the production kinetics, the quantity and quality of the recombinant protein expressed, and the harvest time of the culture. The low and high MOI infections are characterized by their different rate of infection (synchronous Vs. asynchronous) and the associated difference in kinetics of protein expression. Expression of viral vector or virus-like particles, which typically involve co-infecting cell cultures with more than one rBV carrying the coding sequences of individual components adds another complexity to the production process [51], [52]. Altering the MOI of multiple baculoviruses and their time of infection often result in altered capsid compositions as reported by various groups[53], [54]. AAV production using triple baculovirus co-infection strategy is also an example of such a complex and dynamic system where primary work has relied on multicomponent system. Initial studies by Meghrous et al. demonstrated the effect of Bac-Rep on overall Sf9 growth behavior and AAV vector yield using Three-Bac system and its association with encapsidation of vector DNA, and the transducing particle units[48]. The Bac-Rep showed no notable effect on cell viability and growth behavior irrespective of MOI used. The study focused on the multidimensional interactive effect of multiple baculoviruses, each expressing an essential protein of rAAV, suggested a significant effect of Bac-Rep and Bac-Cap MOI on packaging efficiency and functional AAV particles. A full factorial study conducted by Aucoin et al. provided an in-depth analysis of single factor and multifactorial interaction effects of MOI of the three baculoviruses on AAV yields [55]. This study also suggested a self-sustaining nature of insect cell-baculovirus infection process, where differing MOI of one baculovirus did not lead to the drastic changes in overall AAV yield when expressed collectively as relative proportions of total, genome containing and functional particles.