BACKGROUND For several years, orbitally shaken cylindrical bioreactors (OSRs) have become a new approach for propagation of mammalian cells cultivated under suspension. With new commercial cell culture media available today, allowing exceptionally high cell densities, the identification of suitable operating conditions presents a special challenge. Balancing hydrodynamic stress and other factors in reactors, this is particularly true for gas transfer rates providing sufficient oxygen (O-2) to cultures that can attain ten-fold greater biomasses than just a decade ago. This challenge is more profound the larger the working volume of the reactor becomes. RESULT For 10-L working volume cylindrical OSRs, we established a computational fluid dynamics model to evaluate the impacts of different operational conditions. Both fluid velocity distributions and O-2 transfer coefficients (k(L)a) were calculated for different filling volumes and shaking speeds. Also, hydrodynamic stress parameters were investigated considering the most critical working conditions. CONCLUSION The fluid velocity decreased with larger working volumes and, as expected, was found to increase with higher shaking frequency. Gas transfer rates between 10 and 54 h(-1) were found in working volumes from 2.5 to 10 L and under shaking frequencies from 100-160 rpm. Critically assessing the obtained data, with no liquid-submerged gas supply, we conclude that an air flow into the head-space of a disposable bag appears to be satisfactory under certain operating conditions to support the culture of the most sensitive mammalian or insect cell cultures towards high cell density, presently in fed-batch cultures achieving <= 30 x 10(6) cells mL(-1). Shear stress parameters were found to be exceptionally low. (c) 2021 Society of Chemical Industry (SCI).