In this work the maximal operational hydrodynamic conditions (agitation and aeration rate) that cause no adverse effect in Sf-9 cells growth in SF900II serumfree medium were determined. Shear stresses higher than 1 N m -2 and aeration rates higher than 0.04 vvm affect cell growth and when these conditions increase to 1.5 N m -2 and 0.11 vvm, cell growth is completely inhibited with significant cell morphology changes and a strong decrease in viability.
Although the pO 2 did not show a significant effect upon cell growth in the range from 10 to 50%, cell infection and specific productivity were dramatically affected. The production was optimal at a pO 2 of 25% with decreases higher than 50% being observed when the pO 2 decreased to 10 or increased to 50%.
The maximum product quality, i.e., the percentage of product in the form of high molecular weight particles, is not coincident with maximum product titer. Although the highest Pr55gag particle titer was obtained at 96 hours post infection (hpi) and at pO 2 of 25%, the best product quality (defined by gel filtration chromatography and Western immunoblot) was obtained at 48 hpi, independently of the pO 2 used.
The effect of overcritical conditions upon productivity was also studied. As obtained for cell growth, cell infection is affected by shear stresses above 1 N m -2 and by aeration rates higher than 0.04 vvm, with decreases in Pr55gag particle titer higher than 70%, even when the overcritical values are still far from the limit at which cell death occurs.
The results obtained and the optimization strategy used allowed the maximization of the oxygen supply without damaging the cells, with important consequences on the scale-up of a production process involving this insect cell/baculovirus expression system.