On‐line biomass monitoring of CHO perfusion culture with scanning dielectric spectroscopy

Abstract

In this work, dielectric spectroscopy was used to monitor two CHO perfusion culture experiments (B14 and B16). The capacitance of the cell suspension was recorded every 20 minutes over an excitation frequency range of 0.2 MHz to 10.0 MHz. A phase plot of the capacitance at a low excitation frequency vs. the value at a higher frequency proved to be an accurate indicator of the major transition points of the culture, i.e., maximum cell viability, end of lactate consumption, point of zero viability. For both experiments, the capacitance signal correlated very well (R(2) >0.98) with viable cell number up to concentrations of 1 x 10(7) cells/mL. Visual observation of the capacitance spectra indicated that changes in the capacitance relative to frequency were related to the cellular morphology. A multivariate model was developed using off-line data that could predict the median cell diameter within a single experiment (B14) with an error of 0.34 microm (2%). Upon extension to a subsequent experiment (B16), the predicted error was 1.18 microm (9%).

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

“…4 One of the most important parameters to monitor in any cell culture process is the viable, that is, metabolically active biomass. 5,6 Its estimation is almost a philosophic problem. 6 In fact, its most commonly used definition is rather operational and might in some case also include necromass.…”

“…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.…”

“…31 Another study using the same method reports the monitoring of physiological changes and major transition points in CHO cell perfusion culture from multifrequency permittivity data. 5 An alternative to PLS modeling are recently available software packages that allow the automated calculation of the most important b-dispersion parameters. The interpretation of these parameters, like the characteristic frequency (f C ), the permittivity increment (De max ), and others result in additional information on the state of the culture.…”