Biostat b dcu control unit

Cell culture material was generated using an IgG₄ producing cell line (CY01) kindly provided by Lonza Biologics (GB-Slough, Berkshire, UK) in a 5-L (3.5-L working volume) bioreactor with a stirred tank reactor (STR) with an in-built control system (B. Braun BIOSTAT B-DCU control unit, Sartorius, Epsom, UK). Set points were maintained at 30% air dissolved oxygen tension (DO), 7.10 pH and a temperature of 37°C. A constant gas flowrate of 100 cm³/min was maintained using a horseshoe sparger. Agitation was set at 260 RPM using a single 45° pitch, three-blade impeller. All cell cultures were carried out in fed-batch mode using chemically defined medium (CDCHO, Invitrogen, Paisley, UK). The glucose concentration was measured daily with a NOVA Bioanalyser (Nova Biomedical, Deeside, UK) and maintained at a concentration of 2 g/L using a bolus feed of ten-fold concentrated dry powder CD CHO media, adjusted to a concentration of 150 g/L glucose (Sigma, Poole, Dorset, UK).

Cell culture was carried out using the CY01 cell line, kindly donated by Lonza Biologics, in 5 L STRs (B. Braun BIOSTAT B‐DCU control unit, Sartorius, Epsom, UK) and harvested on day 13. Cell culture set points have been carried out as previously described in Popova et al. 27. Total cell concentrations at harvest were ∽9 × 10⁶ cells/mL with an average viability of 77%. The harvested material was used to generate cell culture test materials with the representative most challenging target conditions for future primary recovery feeds. These included as a cell density of 100 × 10⁶ cell/mL with a cell viability of 40%, IgG₁ concentration of 20 g/L and a HCP concentration of 20 g/L. These conditions were selected using a survey compiling expert opinion on the likely future cell culture profiles to primary recovery. Additionally, a low viability was selected to provide a challenging case for the selected technologies. The CCTM generation was described previously in detail 27. The cell culture harvest was concentrated and spiked with the volumes of IgG₁ and a HCP stock to create the required conditions. Apoptosis induced cell stock was added to the CCTM in order to achieve the target viability of 40%. All cell density and viability measurements were carried out using a ViCell™ (by trypan blue exclusion).