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This study described a successful application of the Quality by Design (QbD) approach to pseudorabies virus (PRV) production process development in a fixed-bed bioreactor using the serum-free medium (SFM). The innovated tube-fixed-bed bioreactor was used as a scale-down model of the fixed-bed bioreactor for process development. Risk analysis was performed using Ishikawa diagram combined with failure mode effects analysis (FMEA). The comparative experiment was performed to screen proper medium for adherent African green monkey kidney (Ve