Application of the design of experiment to characterize depth filtration for bispecific antibody clarification

Depth filtration is crucial in the downstream process to clarify the cell culture fluid. With an increase in the upstream cell expression and cell density, achieving a high capacity of clarification (CLR) and the excellent removal of host cell proteins (HCPs) is an enormous challenge. We report a quality-by-design (QbD) case study of clarification screening to help understand the main functional relationships that link the process parameters to product quality or process attributes with prior process knowledge, risk assessment, and the design of experiment (DoE) approach. Our preliminary risk assessment showed that the pore size, charge, and inlet flux of the deep cassette significantly impacted clarification filtration. The DoE of the clarification capacity, protein purity, recovery, and HCP removal was conducted by selecting a two-stage depth filters (primary filters: 4070PC and S870PC; secondary filters: H0HP, H2HP, and H4HP) with different pore sizes, charge differences and inlet flow rates (100 / 200 LMH (L/m²/h)). Depth filter encapsulation and the lower feed flux increased the filtration capacity and HCP removal, while other factors, such as pore size, affected the filtration process time and pressure and were not significant factors in terms of capacity and product quality. Our results suggest that these process parameters can effectively improve the overall performance of the downstream process.