Impact of aeration strategies on fed-batch cell culture kinetics in a single-use 24-well miniature bioreactor

The need to bring new biopharmaceutical products to market more quickly and to reduce final manufacturing costs is driving early stage, small scale bioprocess development. This work describes a comprehensive engineering characterisation of a novel, single-use 24-well parallel miniature bioreactor system. Cell culture performance is also investigated, with particular focus on the aeration strategies adopted at this small scale (7 mL) either by headspace sparging alone or by direct gas sparging into the culture medium. Apparent volumetric oxygen mass transfer coefficient (kLa) values ranged between 3–22 h−1 and 4–53 h−1 for headspace aeration and direct gas sparging respectively. The higher kLa values with direct gas sparging correlated directly with the increase in gas–liquid interfacial area per unit volume. Mixing times (tm) over a range of conditions were generally in the range 1–13 s and decreased with increasing shaking frequency (500–800 rpm). Direct gas sparging also served to reduce tm values by a factor of up to 19 fold. The impact of aeration strategies on cell culture kinetics of a model CHO cell line was also determined. Cultures performed with head space aeration alone showed the highest viable cell density (VCD) (15.2 × 106 cells mL−1), viability retention and antibody titre (1.58 g L−1). These were greater than in conventional shake flask cultures due to the improved control of the μ24 bioreactor system. In all cases the miniature bioreactor managed good control of process parameters such as pH 6.95 ± 0.4, temperature T°C 37 ± 0.4 and DO% 57 ± 32. Cultures performed with direct gas sparging showed a 25–45% reduction in VCD (depending on the aeration strategy used) and a similar reduction in antibody titre. Overall this work shows the successful application of the miniature bioreactor system for industrially relevant fed-batch cultures and highlights the impact of the dispersed gas phase on cell culture performance at the small scale.