Integrated mammalian cell culture and growth measurement using headspace analysis: Experimental and modeling results

Closed-system mammalian cell culture methods have gained prominence due to their potential to minimize contamination risks and support compact experimental designs. However, complete closed-systems often encounter challenges such as pH regulation and oxygen supplementation. This study introduces a novel approach that integrates headspace analysis with mammalian cell culture. The method enables in-situ measurement of CO₂ production by cells, providing quantitative understanding of cell growth. Importantly, we developed mathematical models to elucidate the dynamics of pH changes and oxygen depletion, offering predictive insights for optimizing culture conditions for different cell lines. Using HepG2 cells as a model cell line, the present method agreed well with the reference method (i.e., sulforhodamine B assay) on determining the cell growth curve (R² = 0.98). Furthermore, the method demonstrated good precision, with biological replicates showing relative standard deviations below 7 % across 24-, 36-, and 48-hour culture periods. This integrated approach not only provides solutions for mitigating the limitations of closed-system cell culture but also establishes a framework for high-throughput and efficient applications in biomanufacturing and biomedical research.