Optimization and Application of Serum-Free Medium in Chinese Hamster Ovary (CHO) Cells Culture for Enhanced Production Efficiency

The optimization of serum-free media (SFM) for Chinese Hamster Ovary (CHO) cell culture plays a crucial role in biopharmaceutical production by improving cell growth, viability, and recombinant protein yield. In this study, various SFM formulations, including CD CHO and EX-CELL, were evaluated and compared. An optimized SFM formulation containing insulin (5 µg/mL), transferrin (15 µg/mL), and epidermal growth factor (EGF, 5 ng/mL) significantly enhanced cell density (10.1 × 10⁵ cells/mL), viable cell density (9.5 × 10⁵ cells/mL), and recombinant protein production (475 mg/L) compared to commercial formulations. Growth kinetics analysis showed that CHO cells exhibited exponential growth up to day 5, after which growth slowed due to nutrient depletion. Metabolic profiling revealed peak glucose consumption on day 3 and lactate accumulation by day 5, suggesting a metabolic shift in the stationary phase. The optimized SFM was successfully scaled up to a 5L bioreactor, maintaining high viability (>90%) and consistent protein yield (485 mg/L). Compared to serum-supplemented cultures, the optimized SFM provided comparable protein production with enhanced consistency, scalability, and cost efficiency. These findings underscore the potential of SFM for large-scale biopharmaceutical production, offering an effective and controlled alternative to serum-containing media.