Comparison of a Production Process in a Membrane-Aerated Stirred Tank and up to 1000-L Airlift Bioreactors Using BHK-21 Cells and Chemically Defined Protein-Free Medium

Abstract

The applicability of a protein-free medium for the production of recombinant human interleukin-2 with baby hamster kidney cells in airlift bioreactors was investigated. For this purpose, a BHK-21 cell line, adapted to grow and produce in protein-free SMIF7 medium without forming spheroids in membrane-aerated bubble-free bioreactors, was used as the producer cell line. First, cultivation of the cells was established at a 20-L scale using an internal loop airlift bioreactor system. During the culturing process the medium formulation was optimized according to the specific requirements associated with cultivation of mammalian cells under protein-free conditions in a bubble-aerated system. The effects of the addition of an antifoam agent on growth, viability, productivity, metabolic rates, and release of lactate dehydrogenase were investigated. Although it was possible to establish cultivation and production at a 20-L scale without the use of antifoaming substances, the addition of 0.002% silicon-oil-based antifoaming reagent improved the cultivation system by completely preventing foam formation. This reduced the release of lactate dehydrogenase activity to the level found in bubble-free aerated stirred tank membrane bioreactors and led to a reduction in generation doubling times by about 5 h (17%). Using the optimized medium formulation, cells were cultivated at a 1000-L scale, resulting in a culture performance comparable to the 20-L airlift bioreactor. For comparison, cultivations with protein-containing SMIF7 medium were carried out at 20- and 1000-L scales. The application of protein supplements did not lead to a significant improvement in the cultivation conditions. The results were also compared with experiments performed in a bubble-free aerated stirred tank membrane bioreactor to evaluate the influence of bubbles on the investigated culture parameters. The data implied a higher metabolic activity of the cells in airlift bioreactors with a 150% higher glucose consumption rate. The results of this study clearly demonstrate the applicability of a protein-free chemically defined medium for the production of recombinant proteins with BHK cells in airlift bioreactors.

The role of recombinant proteins in the development of serum-free media

“…Chemically defined media (CDM) using a complex mixture of non-animal derived components (e.g. metals, salts, amino acids) to replace proteins have been achieved for a few cell types (Darfler 1990a;Hesse et al 2003;Spens and Häggström 2005) but growth and production do not always match that of medium containing FCS.…”

Bioreactors: Airlift Reactors

“…The reason for the more successful growth reported in ALRs (5)(6)(7)(8)(9)(10)(11)(12)(13)(14) appears to lie in the difference in the fluid dynamics of ALRs and STRs. In conventional STRs, the energy required for the movement of the fluids is introduced focally at a single point in the reactor, via a stirrer.…”

“…Measurements of fluctuating velocity made show that the liquid turbulence in ALRs is homogeneously distributed in both the riser and the downcomer (31). It thus seems reasonable to assume that the homogeneity of the stress forces is the main advantage offered by ALRs and that this homogeneity is responsible for the success of shear-sensitive cultures in the ALR type of fermenter (5,14,(35)(36)(37)(38)(39)(40). Another advantage of the ALR is the mechanical simplicity of the device.…”

“…Airlift bioreactors are also used for the production of recombinant proteins and monoclonal antibodies from large-scale mammalian cell cultures (14,(437)(438)(439) or to produce penicillin G from immobilized Penicillium chrysogenum in a DT-ALR, utilizing a magnetic field.…”

Membranes in Biotechnology

“…The production of recombinant human interleukin 2 was studied both with and without bubble-free aeration [20]. By modifying the protein-free culture medium SM1F7 by the addition of the surfactant Pluronic F-68 (BASF) the tolerance of the cells towards hydrodynamic stress was increased.…”