On-line monitoring of responses to nutrient feed additions by multi-frequency permittivity measurements in fed-batch cultivations of CHO cells

Ansorge, Sven; Esteban, Geoffrey; Schmid, Georg

doi:10.1007/s10616-010-9267-z

Cited by 46 publications

(42 citation statements)

References 35 publications

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“…Previously, many cell properties have been observed to affect the shape of the beta dispersion, including cell size; intracellular organelle content; membrane‐specific capacitance; and intracellular conductivity . Changes in the shape of the beta dispersion have also been reported for healthy versus nonhealthy cells and for cells undergoing other metabolic changes …”

Section: Introductionmentioning

confidence: 99%

A novel approach for using dielectric spectroscopy to predict viable cell volume (VCV) in early process development

Downey¹,

Graham²,

Breit³

et al. 2014

Biotechnology Progress

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Online monitoring of viable cell volume (VCV) is essential to the development, monitoring, and control of bioprocesses. The commercial availability of steam-sterilizable dielectric-spectroscopy probes has enabled successful adoption of this technology as a key noninvasive method to measure VCV for cell-culture processes. Technological challenges still exist, however. For some cell lines, the technique's accuracy in predicting the VCV from probe-permittivity measurements declines as the viability of the cell culture decreases. To investigate the cause of this decrease in accuracy, divergences in predicted vs. actual VCV measurements were directly related to the shape of dielectric frequency scans collected during a cell culture. The changes in the shape of the beta dispersion, which are associated with changes in cell state, are quantified by applying a novel “area ratio” (AR) metric to frequency-scanning data from the dielectric-spectroscopy probes. The AR metric is then used to relate the shape of the beta dispersion to single-frequency permittivity measurements to accurately predict the offline VCV throughout an entire fed-batch run, regardless of cell state. This work demonstrates the possible feasibility of quantifying the shape of the beta dispersion, determined from frequency-scanning data, for enhanced measurement of VCV in mammalian cell cultures by applying a novel shape-characterization technique. In addition, this work demonstrates the utility of using changes in the shape of the beta dispersion to quantify cell health. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 30:479–487, 2014

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Section: Introductionmentioning

confidence: 99%

A novel approach for using dielectric spectroscopy to predict viable cell volume (VCV) in early process development

Downey¹,

Graham²,

Breit³

et al. 2014

Biotechnology Progress

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“…A sterilizable capacitance probe commercialized as a “biomass monitor” has been available over the last decade. Bulk capacitance and conductance of yeast and mammalian cell cultures can be monitored by a dielectric probe positioned within the bioreactor (Ansorge et al, ; Ducommun et al, ; Knabben et al, ; Opel et al, ; Tibayrenc et al, ). The monitored capacitance of the bulk cell suspension, C , is a function of the effective dielectric permittivity of the suspension, ϵ which is in turn a function of the biovolume.…”

Section: Introductionmentioning

confidence: 99%

The changing dielectric properties of CHO cells can be used to determine early apoptotic events in a bioprocess

Braasch

Nikolic-Jaric

Cabel

et al. 2013

Biotech & Bioengineering

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To ensure maximum productivity of recombinant proteins it is desirable to prolong cell viability during a mammalian cell bioprocess, and therefore important to carefully monitor cell density and viability. In this study, five different and independent methods of monitoring were applied to Chinese hamster ovary (CHO) cells grown in a batch culture in a controlled bioreactor to determine cell density and/or cell viability. They included: a particle counter, trypan blue exclusion (Cedex), an in situ bulk capacitance probe, an off-line fluorescent flow cytometer, and a prototype dielectrophoretic (DEP) cytometer. These various techniques gave similar values during the exponential growth phase. However, beyond the exponential growth phase the viability measurements diverged. Fluorescent flow cytometry with a range of fluorescent markers was used to investigate this divergence and to establish the progress of cell apoptosis: the cell density estimates by the intermediate stage apoptosis assay agreed with those obtained by the bulk capacitance probe and the early stage apoptosis assay viability measurements correlated well with the DEP cytometer. The trypan blue assay showed higher estimates of viable cell density and viability compared to the capacitance probe or the DEP cytometer. The DEP cytometer measures the dielectric properties of individual cells and identified at least two populations of cells, each with a distinct polarizability. As verified by comparison with the Nexin assay, one population was associated with viable (non-apoptotic) cells and the other with apoptotic cells. From the end of the exponential through the stationary and decline stages there was a gradual shift of cell count from the viable into the apoptotic population. However, the two populations maintained their individual dielectric properties throughout this shift. This leads to the conclusion that changes in bulk dielectric properties of cultures might be better modeled as shifts in cells between different dielectric sub-populations, rather than assuming a homogeneous dielectric population. This shows that bulk dielectric probes are sensitive to the early apoptotic changes in cells. DEP cytometry offers a novel and unique technology for analyzing and characterizing mammalian cells based on their dielectric properties, and suggests a potential application of the device as a low-cost, label-free, electronic monitor of physiological changes in cells.

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“…As described in various studies, the cell dielectric properties of cell membrane capacitance ( C M ) and intracellular conductivity (σ i ) could be expressed in two dielectric measures acquired on‐line, the maximal permittivity (Δε max ) and the characteristic frequency of the cell population ( f c ) 27. Dielectric spectroscopy has been increasingly used to monitor a wide range of bioprocesses, from mammalian cell culture to microbiological systems 28–36. Nevertheless, very few studies have described the application of this technology to viral production processes 33, 37, 38.…”

Section: Introductionmentioning

confidence: 99%

Real‐time monitoring of influenza virus production kinetics in HEK293 cell cultures

Petiot

Kamen

2012

Biotechnology Progress

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There is an increased interest from the vaccine industry to use mammalian cell cultures for influenza vaccine manufacturing. Therefore, it became important to study the influenza infection mechanism, the viral–host interaction, and the replication kinetics from a bioprocessing stand point to maximize the influenza viral production yield in cell culture. In the present work, influenza replication kinetics was studied in HEK293 cells. Two infection conditions were evaluated, a low (0.01) and a high multiplicity of infection (1.0). Critical time points of the viral production cycle (infection, protein synthesis, viral assembly and budding, viral release, and host‐cell death) were identified in small‐scale cell cultures. Additionally, cell growth, viability, and viral titers were monitored in the viral production process. The infection state of the cultivated cell population was assessed by influenza immunolabeling throughout the culture period. Influenza virus production kinetics were also on‐line monitored by dielectric spectroscopy and successfully correlated to real‐time capacitance measures. Overall, this work provided insights into the mechanisms associated with the infection of human HEK293 cell line by the influenza virus and demonstrated, once again, the usefulness of multifrequency scanning permittivity for in‐line monitoring and supervision of cell‐based viral production processes. Published 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2013

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On-line monitoring of responses to nutrient feed additions by multi-frequency permittivity measurements in fed-batch cultivations of CHO cells

Cited by 46 publications

References 35 publications

A novel approach for using dielectric spectroscopy to predict viable cell volume (VCV) in early process development

A novel approach for using dielectric spectroscopy to predict viable cell volume (VCV) in early process development

The changing dielectric properties of CHO cells can be used to determine early apoptotic events in a bioprocess

Real‐time monitoring of influenza virus production kinetics in HEK293 cell cultures

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