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.