Parameter Optimization and Capacitance-Based Monitoring of In Situ Cell Detachment in Microcarrier Cultures

This study delves into the scale-down optimization of the in situ cell detachment process for MA 104 cells cultivated on Cytodex 1 microcarriers (MCs). Through a systematic exploration, critical operational parameters—the agitation speed, incubation time, Trypsin–EDTA volume and corresponding activity, and washing steps—were identified as key factors influencing the efficiency and scalability of in situ cell detachment in microcarrier-based cell culture. Maintaining an appropriate agitation speed (1.25 × Njs, minimum agitation speed at which no microcarriers remain stationary for the signification period of 5 s), optimizing the Trypsinization incubation time (up to 60 min), and implementing multiple washing steps (two times) post-medium removal were found to be crucial for efficient cell detachment and subsequent growth. Our study demonstrates the feasibility of reducing the final Trypsin volume to 50 mL per gram of microcarrier while maintaining a Trypsin activity above 380 USP/mL. These conditions ensure complete cell dissociation and improve the cost effectiveness in large-scale productions. Additionally, we introduced real-time monitoring using a capacitance sensor during in situ cell detachment. This method has proven to be an effective process analytical technology (PAT) tool for tracking the cell detachment progress and efficiency. It allows for the prediction of cell detachment based on signals recorded between 3 and 7 min of Trypsinization, enabling rapid process decisions without the need for offline sampling, thereby enhancing the overall process control. This systematic approach not only optimizes in situ cell detachment processes but also has significant implications for the scalability and efficiency of microcarrier-based cell culture systems.