Online determination of viable biomass up to very high cell densities in Arxula adeninivorans fermentations using an impedance signal

Knabben, Ingo; Regestein, Lars; Grumbach, Carsten; Steinbusch, Sven; Kunze, Gotthard; Büchs, Jochen

doi:10.1016/j.jbiotec.2010.06.007

Cited by 16 publications

(6 citation statements)

References 29 publications

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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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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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“…All experiments depicted in this study were executed in a 50 L stirred tank bioreactor (LP351, Bioengineering AG, Wald, Switzerland) described in detail in former publications (Knabben et al, 2010a, b; Knoll et al, 2007; Maier et al, 2001; Regestein et al, 2012). The reactor system was modified to enable calorimetric measurements by determining the heat transfer coefficient UA between fermentation broths and cooling liquid using a calibration heater.…”

Section: Methodsmentioning

confidence: 99%

“…To obtain a constant growth of C. glutamicum during fed‐batch operation under carbon limitation, an exponential feeding was applied. The feeding rate was calculated according to Equation (12) (Knabben et al, 2010a) …”

Section: Methodsmentioning

confidence: 99%

“…Other techniques have been developed to provide more detailed information on the bioprocess, for example exhaust gas analysis, which allows conclusions about the kind of metabolic activity (e.g., respiratory quotient) (Anderlei et al, 2004; Erickson et al, 1978; Gjerkes et al, 2011). Impedance measurement can be used for online determination of viable biomass (Knabben et al, 2010a, 2011; Maskow et al, 2008). For online detection of product formation, however, the particular measuring strategy depends on the specific product.…”

Section: Introductionmentioning

confidence: 99%

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Non‐invasive online detection of microbial lysine formation in stirred tank bioreactors by using calorespirometry

Regestein

Maskow

Tack

et al. 2013

Biotech & Bioengineering

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Non-invasive methods for online monitoring of biotechnological processes without compromising the integrity of the reactor system are very important to generate continuous data. Even though calorimetry has been used in conventional biochemical analysis for decades, it has not yet been specifically applied for online detection of product formation at technical scale. Thus, this article demonstrates a calorespirometric method for online detection of microbial lysine formation in stirred tank bioreactors. The respective heat generation of two bacterial strains, Corynebacterium glutamicum ATCC 13032 (wild-type) and C. glutamicum DM1730 (lysine producer), was compared with the O2 -consumption in order to determine whether lysine was formed. As validation of the proposed calorespirometric method, the online results agreed well with the offline measured data. This study has proven that calorespirometry is a viable non-invasive technique to detect product formation at any time point.

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“…The dielectric properties of biological cells and their different components (cell wall, membranes and cytoplasm) were summarized by Markx and Davey [4]. Over the last couple of decades a lot of research was done on the impedance of cell suspensions, in which a relationship between capacitance and viable cell number was reported [5][6][7][8]. Fehrenbach et al [5] used online capacitive measurements for biomass estimation of Saccaromyces cerevisiae, Pitchia pastoris and Streptomyces virginiae in suspension culture.…”

Section: Introductionmentioning

confidence: 99%

Biomass measurement of living Lumbriculus variegatus with impedance spectroscopy

Sammer¹,

Laarhoven²,

Mejias

et al. 2014

Journal of Electrical Bioimpedance

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Impedance spectroscopy is a useful tool for non-invasive and real time measurements of cell suspensions and a variety of biological tissues. The objective of this study was the investigation of the dielectric properties of living aquatic worms (Lumbriculus variegatus) using impedance spectroscopy in a frequency range between 100 Hz and 10 MHz. We demonstrate a linear relation between the worm biomass and the phase response of the signal thereby providing a quick and precise method to determine the biomass of aquatic worms in situ. Possible applications for nondestructive online biomass monitoring of aquatic worms and other aqueous organisms are discussed. Furthermore, we show that groups of worms fed different diets can be distinguished by the method presented. These results reveal a close relationship between the nutritional composition of the worms and the measured phase response. We also demonstrate that the phase response at 90 kHz does not depend on the worm size. In contrast, the response function for the signal at 440 Hz reveals a linear correlation of average individual worm size and phase. Therefore, we conclude that the measured phase response at 90 kHz qualifies as a measure of the total amount of worm biomass present in the measuring cell, whereas the phase measurement at 440 Hz can be used to estimate the average individual worm size.

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Online determination of viable biomass up to very high cell densities in Arxula adeninivorans fermentations using an impedance signal

Cited by 16 publications

References 29 publications

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

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

Non‐invasive online detection of microbial lysine formation in stirred tank bioreactors by using calorespirometry

Biomass measurement of living Lumbriculus variegatus with impedance spectroscopy

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