Evaluation of media intensification strategies in perfusion.

Upstream continuous manufacturing of biologic therapeutics has been an attractive method to address growing demand. However, maintaining high cell densities (>100 million cells/mL) for long periods of time (>25 days) requires large amounts of media, complicating logistics and presenting a roadblock to increased adoption. To reduce volumetric media use, intensified media recipes have been developed. This study evaluates two such recipes to reduce perfusion rate by up to 30%: the first, a concentrated media formulation to increase nutritional depth, and the second, a commercial perfusion rate reducing supplement. We observe how the cell process, productivity, and product quality are impacted. Direct media intensification stalls cell growth by increasing osmolality, ultimately decreasing cell viability. Meanwhile, media with the perfusion rate reducing supplement maintains cell viability and productivity and has no significant impact on product quality compared to the control. We demonstrated the generalizability of the perfusion rate reducing supplement by testing with a different media and cell clone and observed similar results. Cell cycle analysis demonstrated that both intensified media strategies successfully stall cells in the G0/G1 phase, but the perfusion rate reducing supplement more effectively arrests cell proliferation, suggesting a larger shift from growth-driven cells in glycolysis to production-driven cells in oxidative metabolism, and likely contributing to its superior performance. These findings demonstrate important considerations in media intensification and introduce the perfusion rate reducing supplement as a viable media supplement to reduce volumetric media consumption while maintaining process performance.