Suspension culture titration: A simple method for measuring baculovirus titers.

The baculovirus-insect cell expression system is an important technology for the production of recombinant proteins and baculovirus-based biopesticides. Budded virus titration is critical when scaling up baculovirus production processes in suspension cultures, to ensure reproducible infections, especially when a low multiplicity of infection (MOI) is applied. In this study, a simple suspension culture titration (SCT) assay was developed that involves accurate measurements of the initial cell densities (ICDs) and peak cell densities (PCDs) of an infected culture, from which the MOI and hence the virus inoculum infectious titer can be estimated, using the established Power-Nielsen baculovirus infection model. The SCT assay was assessed in parallel with two adherent culture-based assays (MTT and AlamarBlue) for the Heliothine baculovirus HaSNPV, and was shown to be more objective, time-efficient and reproducible. The model predicted a linear correlation between log(PCD/ICD) and log(MOI), hence an alternative model-independent SCT assay was also developed, which relies on a well-replicated standard curve relating suspension culture-derived PCD/ICD ratios with plaque or endpoint assay-derived MOIs. Standard curves with excellent linearity were generated for HaSNPV and the industrially significant rAcMNPV, demonstrating the feasibility of this simple titration approach, especially in terms of its applicability to a wide range of virus infection kinetics.