Isotope labeling to determine the dynamics of metabolic response in CHO cell perfusion bioreactors using MALDI‐TOF‐MS |
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| Authors: | Marie R. G. Kopp Miroslav Soos Renato Zenobi Massimo Morbidelli |
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| Affiliation: | 1. Institute for Chemical and Bioengineering, Department of Chemistry and Applied Biosciences, ETH Zurich, Zurich, Switzerland;2. Department of Chemical Engineering, University of Chemistry and Technology, Technicka 3, Prague, Czech Republic;3. Laboratory of Organic Chemistry, Department of Chemistry and Applied Biosciences, ETH Zurich, Zurich, Switzerland |
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| Abstract: | The steady‐state operation of Chinese hamster ovary (CHO) cells in perfusion bioreactors requires the equilibration of reactor dynamics and cell metabolism. Accordingly, in this work we investigate the transient cellular response to changes in its environment and their interactions with the bioreactor hydrodynamics. This is done in a benchtop perfusion bioreactor using MALDI‐TOF MS through isotope labeling of complex intracellular nucleotides (ATP, UTP) and nucleotide sugars (UDP‐Hex, UDP‐HexNAc). By switching to a 13C6 glucose containing feed media during constant operation at 20 × 106 cells and a perfusion rate of 1 reactor volume per day, isotopic steady state was studied. A step change to the 13C6 glucose medium in spin tubes allowed the determination of characteristic times for the intracellular turnover of unlabeled metabolites pools,  (≤0.56 days), which were confirmed in the bioreactor. On the other hand, it is shown that the reactor residence time  (1 day) and characteristic time for glucose uptake  (0.33 days), representative of the bioreactor dynamics, delayed the consumption of 13C6 glucose in the bioreactor and thus the intracellular 13C enrichment. The proposed experimental approach allowed the decoupling of bioreactor hydrodynamics and intrinsic dynamics of cell metabolism in response to a change in the cell culture environment. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1630–1639, 2017 |
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| Keywords: | perfusion culture Chinese hamster ovary cells stable isotope labeling MALDI‐TOF‐MS bioreactor dynamics |
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(≤0.56 days), which were confirmed in the bioreactor. On the other hand, it is shown that the reactor residence time 
(1 day) and characteristic time for glucose uptake 
(0.33 days), representative of the bioreactor dynamics, delayed the consumption of 13C6 glucose in the bioreactor and thus the intracellular 13C enrichment. The proposed experimental approach allowed the decoupling of bioreactor hydrodynamics and intrinsic dynamics of cell metabolism in response to a change in the cell culture environment. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1630–1639, 2017