ABSTRACT
Monitoring mammalian cell culture with UVvis spectroscopy has not been widely explored. The aim of this work was to calibrate Partial Least Squares (PLS) models from off-line UVvis spectral data in order to predict some nutrients and metabolites, as well as viable cell concentrations for mammalian cell bioprocess using phenol red in culture medium. The BHK-21 cell line was used as a mammalian cell model. Spectra of samples taken from batches performed at different dissolved oxygen concentrations (10, 30, 50, and 70% air saturation), in two bioreactor configurations and with two strategies to control pH were used to calibrate and validate PLS models. Glutamine, glutamate, glucose, and lactate concentrations were suitably predicted by means of this strategy. Especially for glutamine and glucose concentrations, the prediction error averages were lower than 0.5060.10 mM and 2.2160.16 mM, respectively. These values are comparable with those previously reported using near infrared and Raman spectroscopy in conjunction with PLS. However, viable cell concentration models need to be improved. The present work allows for UVvis at-line sensor development, decrease cost related to nutrients and metabolite quantifications and establishment of fed-batch feeding schemes.
Subject(s)
Bioreactors , Cell Culture Techniques/methods , Cell Proliferation , Spectrophotometry, Ultraviolet/methods , Animals , Calibration , Cell Line , Culture Media/chemistry , Culture Media/metabolism , Least-Squares Analysis , Multivariate Analysis , Oxygen/analysis , Oxygen/metabolism , Sodium Bicarbonate/analysis , Sodium Bicarbonate/metabolismABSTRACT
This work focused on determining the effect of dissolved oxygen concentration (DO) on growth and metabolism of BHK-21 cell line (host cell for recombinant proteins manufacturing and viral vaccines) cultured in two stirred tank bioreactors with different aeration-homogenization systems, as well as pH control mode. BHK-21 cell line adapted to single-cell suspension was cultured in Celligen without aeration cage (rotating gas-sparger) and Bioflo 110, at 10, 30 and 50 % air saturation (impeller for gas dispersion from sparger-ring). The pH was controlled at 7.2 as far as it was possible with gas mixtures. In other runs, at 30 and 50 % (DO) in Bioflo 110, the cells grew at pH controlled with CO2 and NaHCO3 solution. Glucose, lactate, glutamine, and ammonium were quantified by enzymatic methods. Cell concentration, size and specific oxygen consumption were also determined. When NaHCO3 solution was not used, the optimal DOs were 10 and 50 % air saturation for Celligen and Bioflo 110, respectively. In this condition maximum cell concentrations were higher than 4 × 10(6) cell/mL. An increase in maximum cell concentration of 36 % was observed in batch carried out at 30 % air saturation in a classical stirred tank bioreactor (Bioflo 110) with base solution addition. The optimal parameters defined in this work allow for bioprocess developing of viral vaccines, transient protein expression and viral vector for gene therapy based on BHK-21 cell line in two stirred tank bioreactors with different agitation-aeration systems.
