In bioprocess engineering the Qualtiy by Design (QbD) initiative encourages the use of models to define design spaces. However, clear guides on how models for QbD are validated are still missing. In this review we provide a comprehensive overview about validation methods, mathematical approaches and metrics currently applied in bioprocess modeling. The methods cover analytics for data used for modeling, model training and selection, measures for predictiveness and model uncertainties. We point out general issues in model validation and calibration for different types of models and put this into context of existing health authority recommendations. This review provides the start-point for developing a guidance for model validation approaches. There is no one-fits-all approach but this review shall help to identify the best fitting validation method or combination of methods for the specific task and type of bioprocess models that is developed.

Due to their high specificity, monoclonal antibodies (mAbs) have garnered significant attention in recent decades, with advancements in production processes, such as high seeding density (HSD) strategies, contributing to improved titers. This study provides a thorough investigation of high seeding processes for mAb production in Chinese hamster ovary (CHO) cells. Using a metabolic network and flux balance analysis, we compared standard fed-batch (STD FB) with HSD cultivations, exploring supplementary lactate and cysteine, and a bolus medium enriched in amino acids. We observed high glycolytic fluxes and depletion of asparagine simultaneously with the lactate shift building kinetic models around this and other observations. An ensemble of kinetic models assessed the impact of a feeding medium enriched with additional asparagine, revealing a missing lactate shift without enhancing mAb productivity in the experiments. This research provides valuable model-based insights into cellular metabolism during HSD processes, laying the foundation for refined CHO cell-based mAb production. The proposed hypothesis on the regulations guides future experimental validation and bioprocess optimization for enhanced mAb production efficiency.