The purpose of the seed train is to generate an adequate number of cells for the inoculation of the production bioreactor. Adaptygen’s seed train consists of Corning shake flasks, sized 50 (A) and 500 mL (B), 1 L batch spinner bioreactor (C), and a 20 L fed-batch spinner bioreactor (D). The production bioreactor (E) consists of a 1000 L perfusion spinner reactor. The production bioreactor can support high viable cell density (VCD) due to the continuous media exchange rate defined by the cell-specific perfusion rate, the cell bleed to control VCD, and the use of micro and macro spargers to ensure efficient gas exchange occurs within the reactor.

Kinetic models have been utilized to determine the reactor sizes and seed train timeline. The image on the right shows the modeling results of CHO cells for the seed train. Nine production bioreactor runs at a length of 35 days will occur annually in order to reach the targeted mAb yield of 100 kg/year.

(A) Alternating Tangential Flow Separation: Adaptygen’s perfusion system utilizes alternating tangential flow separation (ATF) as a cell retention device for continuous media exchange that allows for the maintenance of the peak viable cell density.

Diafiltration – Tangential Flow Filtration #1: Adaptygen’s first Tangential Flow Filtration (TFF) unit will use a hollow fibre module in a constant volume diafiltration system to primarily desalt the supernatant that is processed from protein A chromatography.

Hollow fibre modules were chosen over cassettes as they offer high packing density, high surface-to-volume ratio, and a smoother and gentler separation. The unit will be 0.415 m long with a surface area of 2.6 m2.

(B) Protein A Capture Chromatography: Protein A chromatography capture of mAbs is the industry standard for primary separation. MabSelect SuRe LX resin was chosen due to its high dynamic binding capacity and superior clearance of HCP. To fully account for the kinetics, a general rate model (GRM) was developed. The GRM works by performing a differential mass balance in the void space and at radial pore positions within a resin bead for each axial position in the column.

(C) Anion Exchange Chromatography: AEX exploits net surface charges to separate impurities from the mobile phase. The AEX column will operate in flow through mode, where impurities bind to the resin while the mAbs pass through the column. The chosen resin for the operation is Nuiva Q due to its high-performance polishing and data availability. The GRM developed for Protein A Chromatography was used again to provide comprehensive modelling of the AEX column.

(D) Multi Modal Cation Exchange Chromatography: MMCEX is the last chromatography column in the downstream operation and completes the final polishing. Nuvia cPrime was selected over other options because it has a sharper breakthrough curve with marginally higher dynamic binding capacities.

(E) Ultrafiltration – Tangential Flow Filtration: The final unit of the downstream process is ultrafiltration through a TFF system. This concentration step involves the supernatant flowing tangentially across a membrane, allowing impurities to flow across the membrane into a permeate stream while mAbs remain in the retentate stream. The model was based on Gel Polarization Model where the bulk concentration of the supernatant is related to the permeate flux and wall or gel concentration