A form of seed train intensification, N-1 perfusion refers to the intensification of cell growth in the step prior to the production bioreactor (N). This process intensification is done by attaching a cell retention device, such as the XCell ATF® Device, to the N-1 bioreactor to attain high cell density and viability. This seeds the production bioreactor at a higher starting cell density and shortens the production bioreactor run time. This can dramatically increase the facility output without direct change to the core production process. A robust cell retention device is required to attain a high cell density inoculum for the production bioreactor.
Start intensification with N-1
The combination of implementation ease and productivity increase make N-1 perfusion the most frequent starting point for upstream intensification. Once in place, further intensify the seed train, add HPH to the N bioreactor or transition to continuous.
N-1 perfusion intensifies the Fed-Batch process, increasing facility throughput. Seeded at a higher initial density, cells in the N bioreactor grow faster, reach a higher max VCD and produce more product than traditional Fed-Batch. Harvest sooner and increase the number of batches per year as compared to a standard fed batch process. Alternatively, maintain the N-bioreactor run time to increase overall yield.
N-1 perfusion intensification increases N bioreactor productivity and throughput while maintaining a fed batch process. No perfusion or media exchange of the N bioreactor is required, allowing the N bioreactor to maintain a single and discreet fed batch harvest point.
Intensify with minimal process modifications and regulatory edits. No additional equipment is connected to the N bioreactor, no media exchange occurs during the production run and cell viability is higher, typically leading to lower impurity values.
N-1 perfused Fed-Batch reaches 350 million cells/mL VCD in just 6 days. In comparison, standard fed-batch achieves 30 million cells/mL viable cell density (VCD) over a course of 14 days (data not shown). Productivity of the N bioreactor after with an N-1 perfusion increased 20-fold (data not shown) while maintain viability above 90%.
Higher cell density and viability
The perfused N-1 bioreactor cell density rapidly increases from 8 to 80 million cells/mL (10X VCD) in 4-5 days. Inoculation of the N bioreactor with the N-1 perfused culture establishes a 10X higher initial viable cell density over fed-batch (A). After 3 days, the N-1 inoculated bioreactor achieves 35 million VCD. In comparison, the control requires 6 days to achieve 25 million VCD (B). Leverage a higher VCD to either increase throughput or increase yield. Harvest of the N-1 Fed- Batch bioreactor at day 8 is equivalent to harvesting the fed-batch control at day 14, saving 6 days of N bioreactor suite time (C). Alternatively, harvest the N-1 fed-batch process at day 14 for a 25% overall yield increase.
Alternating Tangential Flow (ATF) Filtration
An award-winning technology, ATF allows the removal of spent media while keeping cells in culture. Applied using an XCell ATF® device attached to a bioreactor, ATF minimizes cell shear and keeps cells in constant equilibrium with bioreactor contents. This results in faster cell growth at higher densities with higher productivity. Today, the XCell ATF® device, in single-use or reusable format, is the leading perfusion device for mAb and rProtein production.
Xcell ATF® Technology
XCEll ATF® Devices
Potent cell culture supplements
Repligen developed and manufactures cell culture supplements that provide the benefits of serum supplementation while maintaining an animal-free process.
A Brief History of Perfusion Biomanufacturing: How High-Concentration Cultures Will Characterize the Factory of the Future
by John Bonham-Carter and Jerry Shevitz
BioProcess International, October 2011
Developing an Integrated Continuous Bioprocessing Platform: Interview with Konstantin Konstantinov
by Maribel Rios
BioProcess International, December 2012