The scale-up of bioprocesses is still one of the major obstacles in biotechnological industry. Scale-down bioreactors were identified as valuable tools to investigate the heterogeneities observed in large-scale tanks in laboratory-scale. Additionally, computational fluid dynamics (CFD) simulations can be used to gain information about fluid flow in tanks used for production. Here we present the rational design and comprehensive characterization of a scale-down setup, in which a flexible and modular plug-flow reactor is connected to a stirred tank bioreactor. With the help of CFD the mixing time difference between differently scaled bioreactors were evaluated and used as scale-down criterium. Additionally, it was used to characterize the setup at conditions were experiments could technically not be performed. This was the first time a scale-down setup was tested on high cell density Escherichia coli cultivations to produce industrial relevant antigen-binding fragments (Fab). Reduced biomass and product yields were observed during the scale-down cultivations. Additionally, the intracellular Fab fraction was increased by using the setup. The results show that including CFD in the design and characterization of a scale-down reactor can help to keep a connection to production scale and also gain intensive knowledge about the setup, which enhances usability.

Bioprocess development and optimization is a challenging, costly, and time-consuming effort. In this multidisciplinary task, upstream processing (USP) and downstream processing (DSP) are conventionally considered distinct disciplines. This consideration fosters “one-way” optimization without considering interdependencies between unit operations; thus, the full potential of the process chain cannot be achieved. Therefore, it is necessary to fully integrate USP and DSP process development to provide balanced biotechnological production processes. The aim of the present study was to investigate how different host/leader/antigen binding fragment (Fab) combinations in E. coli expression systems influence USP and primary recovery performance and the final product quality. We ran identical fed-batch cultivations with 16 different expression clones to study growth and product formation kinetics, as well as centrifugation efficiency, viscosity, extracellular DNA, and endotoxin content, which are important parameters in DSP. We observed a severe influence on cell growth, product titer, extracellular product, and cell lysis, accompanied by a significant impact on the analyzed parameters of DSP performance. Our results provide the basis for establishing integrated process development considering interdependencies between USP and DSP. These interdependencies need to be understood for rational decision-making and efficient process development.