On‐line determination of animal cell concentration in two industrial high‐density culture processes by dielectric spectroscopy

Abstract

Dielectric spectroscopy was applied to two industrial high cell density culture processes and used to determine on-line the concentration of CHO cells immobilized on macroporous microcarriers in a stirred bioreactor and in a packed-bed of disk carriers. The cell concentration predicted from the spectroscopic data was in excellent agreement with off-line cell counting data for both processes. Deviations between the two counting methods only occurred in the case of a significant decrease of the cell viability, from 93% to 64%, which induced a change of the average cell size in the culture. Results for the packed-bed process were further confirmed by the application of indirect yield models based on the measurement of glucose, lactate, and the protein of interest. Moreover, dielectric spectroscopy was used as a tool to characterize the packed-bed process. It was possible to determine both the maximum cell concentration that could be reached in the culture system, 2.0 x 10(11) cell per kg of disk carrier, and to quantify the increase of specific protein productivity induced by the production phase, from 5.14 x 10(-8) microg x cell(-1) x h(-1) to 4.24 x 10(-7) microg x cell(-1) x h(-1).

Process control in cell culture technology using dielectric spectroscopy

“…Until 2002, most methods for the monitoring of cell concentrations were not applicable to immobilized cell cultures using microcarriers or other fixed bed carriers. Common methods now allowing the monitoring of these cultures include some optical techniques (see above) based on light absorbance and/or scattering, real-time imaging, particle size analysis and measurements of culture fluid density [Ducommun et al, 2002a].…”

Multifrequency permittivity measurements enable on‐line monitoring of changes in intracellular conductivity due to nutrient limitations during batch cultivations of CHO cells

“…The successful application of this technology in a wide variety of cell culture processes is described in the literature. 5,6,9,[13][14][15][16] Several studies report measurements of the permittivity at one single or two frequencies (to reduce the effect of a changing media composition on the signal). Permittivity measurements are described as a tool to directly measure the viable biomass.…”

“…However, this relationship does not always hold because stained cells are known to contribute to the permittivity. 5,13,[17][18][19] This observation is probably caused by the fact that the ''remaining'' permittivity of a nonviable cell is dependent on the mechanism by which the cell was ''inactivated.'' 20 The classical method of counting viable cells by dye exclusion can therefore not always be expected to give data that correlate linearly with the permittivity, in particular in the late stage of cultivations during which viability is low.…”

“…21 Permittivity measurements do also reflect changes in cell physiology which has been demonstrated for a wide variety of cell types. 5,9,13,[22][23][24] Cell physiology is most commonly described by the term ''physiological state.'' This term is defined by several variables that impart significant information about the cellular state.…”

Development of a small-scale bioreactor: Application to in vivo NMR measurement

“…In addition, the smallscale bioreactor has to contain enough cells to be capable of performing noninvasive in vivo as well as ex vivo assays. Current noninvasive in vivo assays or those under development are based on highly sophisticated analytical equipment, such as optical sensors for UV, IR, Raman, and fluorescence spectroscopy, in situ microscopy (Ulber et al, 2003), dielectric spectroscopy (Ducommun et al, 2002), and in vivo nuclear magnetic resonance (NMR) (Castro et al, 1999). These different analytical techniques are highly complementary to obtain an adequate description of cell population physiological, nutritional, and metabolic states and dynamic behaviors.…”