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Upstream process intensification using perfusion cell culture has become a necessity for meeting today’s demand in monoclonal antibodies (mAbs) bioprocessing. By sustaining higher viable cell densities, extending culture duration, and enabling continuous product harvest, perfusion cell culture allows manufacturers to increase volumetric productivity while reducing cost of goods. The demand for scalable production technologies continues to rise as mAbs now account for roughly 60% of biopharmaceutical revenue, with most new approvals aimed at oncology and autoimmune diseases.
Successful scale-up of perfusion cell culture to commercial manufacturing depends on developing a robust, well-characterized process at small scale. Scientists at the Zurich University of Applied Sciences (ZHAW) demonstrated how large perfusion systems can be replicated at 250 mL scale using a Sartorius Ambr® 250 Modular bioreactor, which was not designed for perfusion process intensification, integrated with Repligen’s XCell® ATF 1 (Alternating Tangential Flow) device. Their results position small-scale, fully controlled bioreactors as powerful tools for optimizing clones, media, and perfusion parameters in parallel.
Case studies at 250 mL scale
ZHAW researchers evaluated two perfusion strategies at the 250 mL scale using an Ambr 250 Modular bioreactor coupled to an XCell ATF 1 device. They adapted the silicone lid to enable direct integration of the single-use ATF device into the reactor, thus creating a setup that could be sterilized and operated under standard lab conditions.
N-1 Perfusion:
The first study focused on intensifying seed train expansion. Cultures of antibody-producing CHO cells reached peak densities of approximately 1.7 × 10⁸ cells/mL within seven to eight days, levels that are suitable for inoculating production-scale reactors. The cell quality remained comparable to reference cultures run in a 3 L bioreactor equipped with an XCell ATF 2 device, validating the scalability of the miniature system. Slightly slower growth in the Ambr 250 was attributed to higher agitation speeds required to maintain oxygenation at the smaller volume, which may have introduced shear-related stress.
Continuous Perfusion:
A second proof-of-concept study from ZHAW scientists focused on continuous perfusion. Over a 23-day run, their setup maintained steady-state conditions at target cell volumes and achieved volumetric productivity of approximately 0.65 g/L/day. Even with manual bleed control, the performance of the small-scale set-up closely mirrored that of a larger automated 3 L bioreactor. Furthermore, product quality parameters such as IgG titers and cell-specific productivity were all within expected ranges.
Taken together, these studies demonstrate that it is possible to establish perfusion processes in one of the smallest fully controlled reactor formats available. By integrating the XCell ATF 1 device with the Ambr 250 Modular, ZHAW researchers showed that small-scale systems can reliably match large-scale setups.
Linear scalability matters
Linear scalability is central to process reliability and regulatory confidence. For perfusion cell culture, successful scale-up depends on maintaining the same operating principles and physical conditions across every reactor volume. In practice, that means keeping four key parameters constant: flux rate, backflush ratio, shear, and residence time.
Repligen’s XCell ATF System was engineered with linear scalability in mind. The system uses a diaphragm pump to alternate the flow of the cell culture fluid across a hollow-fiber membrane, creating a backflush effect that continuously removes debris and maintains run longevity.
The system also equalizes the flux rate, ensuring that each filter size processes the same volume throughput, and equalizes flow per fiber across device sizes so that cells experience the same shear environment at all scales. Finally, the system maintains a constant ratio of displacement volume to hold-up volume, keeping residence time and media exchange comparable from bench to production scale. This mechanism is identical across the ATF family of products ensuring that cell performance and mass-transfer dynamics remain comparable at every scale.
From bench to biomanufacturing
Successfully operating perfusion at the 250 mL scale represents a practical advance in process development. In this configuration, the Ambr 250 and the XCell ATF 1 work in tandem to enable parallel screening of multiple clones, media formulations, and feed strategies. Furthermore, because the ATF 1 maintains the same shear and residence-time profile as larger devices, data generated at the bench scale can inform processes at 3–50 L pilot and multi-thousand-liter production volumes. This scalability continuum allows manufacturers to make data-driven decisions earlier, reducing the number of scale-up iterations and facilitating smoother technology transfer.
Supporting sustainability
Whether adjusting perfusion rates or selecting filtration technology, mAb manufacturers must ensure that the decisions they make at the bench translate reliably to production. The ZHAW case studies demonstrate that small-scale systems can match commercial-scale performance and accelerate upstream development. But beyond higher productivity, perfusion also supports the biopharma industry’s sustainability goals. Smaller, more efficient reactors reduce energy, water, and raw material use, minimizing waste production. These capabilities will remain central for maintaining both overall throughput and environmental responsibility as biopharma’s antibody portfolios grow.
