60 BIOTECHNOLOGY
detected by the control station software, and calibration was performed. Unlike an analogue sensor, which stores calibration data only in the control software, the calibration data is stored in the ISM pH sensor itself, allowing it to be recalled at any time. In addition, the sensor can be connected to the optional Mettler-Toledo iSense software via USB. Using this software, a wide range of data is available including the calibration data performed in preparation for this experiment as well as intelligent monitoring of a sensor’s remaining shelf life (sensor ‘health’).
Reliable monitoring and gassing control Both the three-gas and four- gas automatic DO control algorithms allowed the culture to reach similarly high viable cell densities. Te four-gas experiment reached its peak cell density (8.89 x 106
cells/mL)
sooner than the three-gas run (9.54 x 106
consumption and lactate and ammonia accumulation were comparable between the two cultures (Fig.1).
Consistent with the cell
density trend, the three-gas culture consumed glucose slightly slower than the four-gas culture. When the glucose was exhausted, the cell growth and viability began to drop. Higher peak densities would have been possible if glucose and other necessary nutrients had been supplemented using a fed-batch protocol. Te two gassing control algorithms produced comparably healthy cultures, and showed some notable gas consumption differences. Te four-gas culture consumed more gas overall, as illustrated in Fig.1, D. Since the three-gas algorithm does not utilise N2
for DO control,
there is a possibility for the DO to climb above setpoint at the beginning and end of the run when O2
demand is low. Using
four-gas control, N2
www.scientistlive.com
is available to cells/mL). Glucose
keep DO at setpoint, which may be beneficial for some sensitive cell types, and for anaerobic cultures. Whether a culture will be healthier with three-gas or four- gas automatic gassing control will have to be determined empirically for each cell strain. Te evo 200 capacitance biomass sensor was a valuable in-line measure of cell growth during the runs. Tere was also a comparison made between the offline viable cell density measurement and the in-line evo 200 capacitance measurement for one run. After calibrating this sensor for a particular cell line and specific culture process, it can be used in place of sampling the bioreactor, which would avoid lost volume and reduce the risk of contamination. Tree DO sensors were incorporated into these experiments. Te two ISM sensors were automatically detected by the control station, and including the traditional polarographic sensor, all three were able to accurately track and trend DO levels throughout the run. No significant differences were seen between DO measurement by the three sensors. With the use of intelligent sensors, the BioFlo 320 provides advanced process control for
CHO cell culture. Tis method provided similar results using either the three-gas or four-gas automatic gassing cascades. Te setup can be used to meet a host of culture requirements and the upfront knowledge of an ISM sensor’s ‘health’ greatly reduces operational risk due to potential sensor failure during a cell culture run.
Lessons learned as a result of these experiments In these experiments, the ability to customise the configuration by adding an evo 200 biomass sensor and multiple ISM DO sensors elevated these runs to ‘intelligent’ CHO cell culture. With the addition of an in-line bioanalyser, sampling of the bioreactor could be eliminated to reduce the risk of sampling- associated contamination, making the BioFlo 320 a superior setup for cell culture and an intelligent choice for bioprocess laboratories worldwide.
For more information ✔ at
www.scientistlive.com/eurolab
Christiane Schlottbom is with Eppendorf in Europe. Stacey Willard & Ma Sha are with Eppendorf, Inc. in the USA.
www.eppendorf.com
Caption: Fig. 1. Comparing (A) glucose, (B) lactate, (C) ammonia concentrations, and (D) the total gas consumption between three-gas and four- gas during the bioreactor runs
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