Analysis of cell lysis for improved understanding between the shake tube and stirred tank reactor perfusion CHO cell cultures

Shake tubes (ST) are widely employed to assist the development of the stirred tank reactor (STR) perfusion cell culture. However, cell lysis may be frequently underrestimated and lead to culture performance discrepency between these systems, rendering the ST model ineffective in designing the STR perfusion cultures. In this study, perfusion culture performance bewteen the STR and ST was investigated under various conditions with the analysis of cell lysis. Comparable performance was observed bewteen the two systems at low perfusion rates ( $D$ ≤1.0 VVD), except that the specific productivity ( $q_P$ ) of the STR was decreased at $D$ =0.5 VVD, which was related to product degradation by cell lysis. In contrast, significant differences in cell maintenance, metabolism, and $q_P$ were found at $D$ =2.0 VVD. By the analysis of the authentic cell growth and death kinetics, it was found that cell growth arrest, potentially due to the limited availability of oxygen, led to the stable cell maintenance at VCD≈90 × 10⁶ cells/ml and altered cellular metabolism for the ST, while the continuous decline of VCD and $q_P$ in the STR were related to excessive cell death, subsequently ascribed to the harmful hydrodynamic stress conditions. We further demonstrated that cell lysis accounted for 57.62-76.29% of the total generated biomass in both the reactors and significantly impacted the estimation of process descriptors crucial for understanding the true cellular states. With cell lysis in sight, cell performance can therefore be accurately described and this knowledge can be further leveraged to expedite process development for the perfusion cell culture processes.