Multi-wavelength fluorescence spectroscopy was investigated as a potential tool for use in monitoring key process variables that include: viable and dead cells, recombinant protein, glucose, and ammonia concentrations for Chinese hamster ovary (CHO) cells during cultivation.For the purpose of calibrating the fluorescence-based empirical model, cells were grown in batch mode with different initial glucose and glutamine concentrations.Spectrofluorometer settings were optimized to ensure reproducibility and accuracy of the acquired spectra. With the purpose of gaining qualitative insight into the evolution of the spectra, the trajectories of individual fluorophore peaks were studied during the cultivation process. Spectral changes related to biomass and secreted proteins were investigated by comparing the spectra at various stages during the downstream processing. A partial least square regression (PLSR) was used to formulate empirical models that related the input data set, i.e., the fluorescence excitation-emission matrix, to the actual state of the system including viable cell and dead cells and recombinant protein, glucose, and ammonia concentrations. The models exhibited accurate prediction ability for the process variables of interest.
The impact of cell culture environment on the glycan distribution of a monoclonal antibody (mAb) has been investigated through a combination of experiments and modeling. A newly developed CHO DUXB cell line was cultivated at two levels of initial Glutamine (Gln) concentrations (0, 4 mM) and incubation temperatures of (33 and 37 °C) in batch operation mode. Hypothermia was applied either through the entire culture duration or only during the post-exponential phase. Beyond reducing cell growth and increasing productivity, hypothermia significantly altered the galactosylation index profiles as compared to control conditions. A novel semi-empirical dynamic model was proposed for elucidating the connections between the extracellular cell culture conditions to galactosylation index. The developed model is based on a simplified balance of nucleotides sugars and on the correlation between sugars' levels to the galactosylation index (GI). The model predictions were found to be in a good agreement with the experimental data. The proposed empirical model is expected to be useful for controlling the glycoprofiles by manipulating culture conditions.
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