Dissolved carbon dioxide accumulation in a large scale and high density production of TGF? receptor with baculovirus infected Sf-9 cells

Abstract

Production of a TGFβ receptor with high density baculovirus infected Sf-9 cells (7×10(6)cells ml(-1)) served as a test run for a retrofitted 150 L microbial fermentor. The entire 110 L batch run was performed in serum free medium, with an addition of a concentrated amino acid and yeastolate mixture at the time of infection. This addition strategy has been proven effective at a small scale by enabling cultures to maintain maximum product yield. In the bioreactor however, while cellular growth was comparable to that of the smaller scale control, TGFβ receptor production was three fold below the control. To minimize the mechanical stress, low flow rate of pure oxygen was used to control the dissolved oxygen at 40%. As a consequence, it seems that this aeration strategy involved an accumulation of dissolved carbon dioxide that in turn inhibited the protein production. A model has been developed that estimated the CO(2) partial pressure in the culture to be in the vicinity of 0.15 atm. The effect of dissolved CO(2) at this concentration has been assessed at smaller scale for TGFβ receptor and β-gal expression, in controlled atmosphere incubators.

Monitoring lentiviral vector production kinetics using online permittivity measurements

“…This value was used as an indicator of the volumetric oxygen consumption. The overall aeration strategy was similar to what has been described earlier by our group [34].…”

Reactor Engineering in Large Scale Animal Cell Culture

“…If a low enriched air flow rate or pure oxygen are used to enhance the driving force and thereby reduce the flow rate and potential for damage due to bursting bubbles, high levels of pCO 2 of 150-200 mm Hg are predicted to be reached (Gray et al 1996) and such levels have been measured in 1800-2500 L bioreactors and a high cell density perfusion bioreactor (Gray et al 1996;de Zengotita et al 2002). Detrimental effects have been reported for growth and productivity of a recombinant myeloma process (Aunins et al 1993), hybridoma, NSO, BHK, (de Zengotita et al 2002, CHO (Gray et al 1996;Mostafa and Gu 2003) and insect cells (Garnier et al 1996) at these high levels of pCO 2 , though these cells grew well in the rangẽ 35 to~80 mm Hg pCO 2 , comparable to the physiological range of 31-54 mm Hg pCO 2 (de Zengotita et al 2002).…”

“…Overall, Mostafa and Gu (2003) reported an improved productivity (40%), culture time (the fed-batch time was extended from 10 to 14 days) and final titre (2-fold, equivalent to that at the 1.5 L scale). However, they also noted that at constant pH, higher pCO 2 is also associated with higher osmolality and that therefore the improved performance at lower pCO 2 might also be due to reduced osmolality (Garnier et al 1996;de Zengotita et al 2002) but it was not monitored. In passing, it should be mentioned that the use of antifoam enabled the use of higher air flow rates (0.004 vvm) with the sparge stone without excessive foaming and again enabled the culture duration to be extended from 10 to 14 days (Mostafa and Gu 2003).…”

Cell Culture Processes in Monoclonal Antibody Production

“…Kunkel et al reported that reduction in dissolved oxygen caused decreased glycosylation of antibody N-glycan chains. 65 Dissolved CO 2 is also an important process variable because it can accumulate to inhibitory levels at values greater than 120-150 mm Hg [66][67][68] and affects product quality. Even though dissolved CO 2 sensors are commercially available, e.g., from YSI Inc., (Yellow Springs, OH) or Metler Toledo (Columbus, OH), dissolved CO 2 is still measured using off-line blood gas analyzers such as those from Nova Biomedical (Waltham, MA).…”