Accordingly, mixing is of great importance at high cell density cultures ( Al-Rubeai, 2015). Additionally, the combination of jet flow and stirring might induce a direct connection between perfusion feed and retentate flow leading to the formation of a short circuit flow. Besides a technical complex setup, which increases the risk of contamination, high cell densities are responsible for elevated viscosities in the bioreactor and consequently a decrease in mixing efficiency ( Ozturk, 1996). On the downside perfusion mode also comes with challenges. With such intensified fed batch strategies, the titer was almost doubled compared to low seeded cultures ( Stepper et al., 2020). As an alternative to N-stage perfusion or traditional fed batch processes the less-productive growth phase can also be shifted to the pre-stage, leading to higher volumetric productivities in the N-stage. Most common technologies like tangential flow filtration (TFF) or alternating tangential flow (ATF) systems involve a hollow fiber, which is used to circle cells back into the bioreactor while constantly exchanging media ( Karst et al., 2016). Therefore, perfusion processes have become a substantial part of upcoming manufacturing technologies using cell retention to provide higher cell densities. Nevertheless, the need for higher product yields has encouraged the development of intensified process strategies. The majority of mAbs are typically manufactured in Chinese Hamster Ovary (CHO) cells with fed batch processes as preferred process strategy ( Jain and Kumar, 2008 Orellana et al., 2015 Dhara et al., 2018 Kesik-Brodacka, 2018). In particular the demand for monoclonal antibodies (mAbs) has increased over the past years. The results demonstrate how computational fluid dynamic models can be used for rational process design of intensified production processes in the biopharmaceutical industry.Ī rapidly growing population, increased prevalence of diseases as well as rising knowledge and acceptance of biopharmaceuticals has compelled biopharmaceutical companies to optimized processes. Nevertheless, since Emix was confirmed to be >90% also for both 2 L setups and the determined mixing times were in a similar range for all scales, the 2 L system was deemed to be a suitable scale down model. Two different setups were evaluated in 2 L scale where the direction of flow was changed, yielding a difference in mixing efficiency of 10%. No geometric adaptations in the pilot scale systems were necessary as Emix was greater than 90% for all conditions tested. This evaluation gives insight into the flow pattern, the mixing behavior and information on cell residence time inside the bioreactors. Highly resolved Lattice Boltzmann Large Eddy simulations were performed in single phase and mixing efficiencies (Emix) furthermore experimentally validated in the 2 L system. This study investigates the scale up from a 2 L glass bioreactor to 100 L and 500 L disposable pilot scale systems. Computational fluid dynamics can be used to identify such short circuit flows, assess mixing efficiencies, and eventually adapt the perfusion setup. As cells in continuous processes circulate outside the bioreactor, performance losses may arise if jet flow and stirring cause a direct connection between perfusion feed and return. The robust scale up of perfusion systems requires comparable conditions over all scales to ensure equivalent cell culture performance. 3Cell Culture, Process Science, Boehringer Ingelheim, Fremont, CA, United States.2M-Star Center Europe GmbH, Sargstedt, Germany.1Late Stage USP Development, Bioprocess Development Biologicals, Boehringer Ingelheim Pharma GmbH & Co. Maike Kuschel 1 Johannes Wutz 2 Mustafa Salli 1 Dominique Monteil 3 Thomas Wucherpfennig 1*
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