Abstract
An 11‐L helical ribbon impeller (HRI) bioreactor was tested for the culture of Spodoptera frugiperda (Sf−9) cells. This impeller and surface baffling ensured homogeneous mixing and high oxygen transfer through surface aeration and surface‐induced babble generation. Serum‐supplemented and serum‐free cultures, using TNMFH and IPL/41 media, respectively, grew a similar specific growth rates(0.031 and 0.028 h^−1^) to maximum cell densities of 5.5 × 10^6^−6.0 × 10^6^ cells. mL^−1^ with viability exceeding 98% during exponential growth phase. Growth limitation coincided with glucose and glutamine depletion and production of significant amounts of alanine. The bioreactor was further tested under more stringent conditions by infecting a serum‐free medium culture with a recombinant baculovirus. Heterologous protein production of ∼35 μg per 10^6^ cells was comparable to yields obtained in serum‐free cultures grown in spinner flasks and petri dishes. Average specific oxygen up‐take and carbon dioxide production rates of the serum‐free culture prior to infection as measured by on‐line mass spectroscopy were 0.20 μmol O~2~μ·(10^6^ cells)^−1^ h^−1^ and 0.22 μmol CO~2~ · (10^6^ cells)^−1^h^−1^ and increased by 30–40% during infection. Therefore, the mixing and oxygenation conditions of this bioreactor were suitable for insect cell culture and recombinant protein production, with limitation being mainly attributed to nutrient depletion and toxic by‐product generation.