Time-Resolved Monitoring of the Oxygen Transfer Rate of Chinese Hamster Ovary Cells Provides Insights Into Culture Behavior in Shake Flasks

Abstract

Cultivations of mammalian cells are routinely conducted in shake flasks. In contrast to instrumented bioreactors, reliable options for non-invasive, time-resolved monitoring of the culture status in shake flasks are lacking. The Respiration Activity Monitoring Respiration Activity Monitoring System system was used to determine the oxygen transfer rate (OTR) in shake flasks. It was proven that the OTR could be regarded as equal to the oxygen uptake rate as the change of the dissolved oxygen concentration in the liquid phase over time was negligibly small. Thus, monitoring the oxygen transfer rate (OTR) was used to increase the information content from shake flask experiments. The OTR of a Chinese hamster ovary cell line was monitored by applying electrochemical sensors. Glass flasks stoppered with cotton plugs and polycarbonate flasks stoppered with vent-caps were compared in terms of mass transfer characteristics and culture behavior. Similar mass transfer resistances were determined for both sterile closures. The OTR was found to be well reproducible within one experiment (standard deviation <10%). It correlated with changes in cell viability and depletion of carbon sources, thus, giving more profound insights into the cultivation process. Culture behavior in glass and polycarbonate flasks was identical. Monitoring of the OTR was applied to a second culture medium. Media differed in the maximum OTR reached during cultivation and in the time when all carbon sources were depleted. By applying non-invasive, parallelized, time-resolved monitoring of the OTR, the information content and amount of data from shake flask experiments was significantly increased compared to manual sampling and offline analysis. The potential of the technology for early-stage process development was demonstrated.

Non‐invasive and time‐resolved measurement of the respiration activity of Chinese hamster ovary cells enables prediction of key culture parameters in shake flasks

“…As the change of the oxygen concentration in the liquid can be assumed to be very small compared to OUR and OTR, the measured OTR directly reflects the OUR. [29,30] The validity of this assumption for CHO cultures is discussed in detail elsewhere. [29] As demonstrated recently by monitoring the OTR for an industrial CHO cell line, measurement times can be adapted flexibly to allow for the detection of very small changes in the oxygen partial pressure.…”

“…[29,30] The validity of this assumption for CHO cultures is discussed in detail elsewhere. [29] As demonstrated recently by monitoring the OTR for an industrial CHO cell line, measurement times can be adapted flexibly to allow for the detection of very small changes in the oxygen partial pressure. [29] RAMOS also enables monitoring of the carbon dioxide transfer rate (CTR).…”

“…[29] As demonstrated recently by monitoring the OTR for an industrial CHO cell line, measurement times can be adapted flexibly to allow for the detection of very small changes in the oxygen partial pressure. [29] RAMOS also enables monitoring of the carbon dioxide transfer rate (CTR). [28] Here, the increase of the carbon dioxide partial pressure in the gas phase is used.…”

“…This is analogue to the assumption that the OTR will equal the OUR, if the oxygen concentration in the liquid does not change during measurement. [29] Only under this assumption, CER from cell respiration will equal the CTR. From the ratio of OTR and CTR the transfer quotient (TQ) can be calculated.…”

“…Adjustment of RAMOS for utilization of CHO cultures was described recently. [29] As the TOM device was integrated into an incubator where flasks were run in parallel for manual sampling, measurement values were affected by opening the incubator hood for sampling. As a result, OTR and CTR measurement values were correlated with sampling points and values that deviated more than 30% from the previous measurement value were considered outliers and excluded.…”