In this study, we demonstrated the first, to our knowledge, integrated continuous bioprocess (ICB) designed for the production of acid‐sensitive monoclonal antibodies, prone to aggregate at low pH, on pilot scale. A high cell density perfusion culture, stably maintained at 100 × 10⁶ cells/ml, was integrated with the downstream process, consisting of a capture step with the recently developed Protein A ligand, Z_Ca; a solvent/detergent‐based virus inactivation; and two ion‐exchange chromatography steps. The use of a mild pH in the downstream process makes this ICB suitable for the purification of acid‐sensitive monoclonal antibodies. Integration and automation of the downstream process were achieved using the Orbit software, and the same equipment and control system were used in initial small‐scale trials and the pilot‐scale downstream process. High recovery yields of around 90% and a productivity close to 1 g purified antibody/L/day were achieved, with a stable glycosylation pattern and efficient removal of impurities, such as host cell proteins and DNA. Finally, negligible levels of antibody aggregates were detected owing to the mild conditions used throughout the process. The present work paves the way for future industrial‐scale integrated continuous biomanufacturing of all types of antibodies, regardless of acid stability.

Disperse dyes, which are used for colouring synthetic textile fibres, are well-known contact sensitisers. To investigate the outcome of patc...

The production of biopharmaceuticals is a chemical- and water-intensive process. The consumption of water and chemicals is partly due to the need for many different buffers in large volumes during the downstream process, typically consisting of several chromatography steps. Given the global commitment to the goals for sustainable development and the anticipated growth of the biopharmaceutical market, the consumption of large buffer volumes is expected to become problematic. To address this, we propose the introduction of buffer recycling to reduce the consumption of water and chemicals. For solvent based pharmaceutical processes, solvent recycling through re-distillation is already established, but for water-based processes, this concept is still rather unexplored. In this study, buffer recycling was implemented during the equilibration phase of Protein A chromatography for antibody purification. We have investigated the potential gains of buffer recycling and demonstrated buffer recycling in two technical implementations: buffer recycling in a batch-to-batch process and buffer recycling in a multi-column process. Buffer recycling consists of buffer recovery and buffer reuse. During recovery, buffer that has been used during equilibration is collected and pH adjusted, and during the reuse step, the recovered buffer is reintroduced into the process. By introducing buffer recycling, we could reduce the equilibration buffer consumption by almost 50 % in this phase, corresponding to more than 10 % of the total buffer use in the Protein A protocol, and have seen no changes in antibody yield or purity. Hence, through buffer recycling, we can improve the sustainability in biomanufacturing by using less water and chemicals.

The transition from batch-wise to integrated continuous manufacturing of biopharmaceuticals has gained significant attention over the past decade. While this concept has been thoroughly studied, there remains significant room for improvement in terms of process automation and real-time monitoring. This study showcases the implementation of an integrated downstream process at a lab scale, operating continuously and autonomously. The purification process included solvent/detergent-mediated virus inactivation in a packed bed reactor, Protein A capture via a three-column periodic counter-current chromatography process with UV-based loading, and a mixed-mode chromatography polishing step in flow-through mode. Support systems for at-line automated sample collection and high-performance-liquid chromatography analysis for near-real-time quality monitoring, along with autonomous buffer management, were introduced. The process operated for over five days, with two disturbances introduced: decreased capture column capacity and increased harvest titer. The high level of automation and control allowed the process to adapt to these disturbances. Throughout the run, the product was sampled, and quality analyses were performed, achieving an average monomer content of >98.5 % and an overall yield >90 %. This study demonstrates the feasibility of introducing autonomous integrated continuous downstream processes at a lab scale, enabling users to concentrate on process and control development.

The downstream process of monoclonal antibodies (mAbs) is expensive and significantly contributes to overall manufacturing costs. One primary reason is the extensive consumption of water and chemicals required for preparing large volumes of various buffers, essential for multiple chromatography and filtration steps. Reducing the water consumption in biopharmaceutical processes is critical to drive down costs and improve sustainability, which can be achieved through the introduction of buffer recycling. In this study, we implemented buffer recycling in an integrated two-step mAb downstream process consisting of a Protein A capture step in a periodic counter-current (PCC) set-up, followed by a mixed-mode polishing step in flowthrough mode. Buffer recycling was implemented during the cleaning-in-place (CIP) phases of the integrated steps, where the CIP buffer from the polishing column was recovered and reused counter-currently in the CIP phase of the capture column. Compared to the reference process without buffer recycling, this approach resulted in 29% savings in CIP buffer, while maintaining product purity within 0.66% and yield within 1.68% of the reference process. These minor differences confirm that buffer recycling can be implemented without compromising product quality. Through buffer recycling, we see significant potential to improve process sustainability in biomanufacturing by conserving water and reducing chemical waste.