In mammalian cell culture, single-use bioreactors are widely used. Different hardware designs are available, ranging from stirred tank reactors to wave mixed and cubical shaken systems. Unlike in stainless steel systems, where standards exist, in singleuse bioreactors aeration devices are often predefined by the supplier. While ring sparger systems are the gold standard in stainless steel bioreactors, not all singleuse bioreactors are available with ring spargers. In this study, a comprehensive characterization of two stirred tank single-use bioreactor systems (XDR TM from Xcellerex and S.U.B. from Thermo Scientific Hyclone) was performed under GMP conditions with 200/250 L and 1000 L bioreactors. Engineering facts like mass transfer rates for oxygen k L a o2 and carbon dioxide k L a CO2 as well as mixing number were evaluated. To achieve improved similarity in key engineering parameters and in consequence cell culture performance, the submerse aeration device of the S.U.B.(to date only open tube and frit) was remodeled resulting in a drilled hole sparger design. Results of the characterization showed that k L a o2 in the S.U.B. was enhanced from 8.5 h −1 to 11.5 h −1 at the maximum, and the k L a CO2 was very similar between both bioreactor types. Knowledge of the characterization data as well as improved oxygen transfer rate in the S.U.B. allows for an interchangeable usage of the two different single-use bioreactors.