On the audio- and radio-frequency dielectric behaviour of anchorage-independent, mouse L929-derived LS fibroblasts

Davey, Christopher L.; Kell, Douglas B.; Кемп, Р. Б.; Meredith, R. W. J.

doi:10.1016/s0302-4598(98)80007-x

Cited by 46 publications

(9 citation statements)

References 42 publications

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“…The decrease of the mean C, value (and its CV) in hypotonically swollen cells is due to the disappearance of membrane folds and microvilli (12,19,37,38). At physiological osmolality (280-300 mOsm), large variations in the plasma membrane morphology cause the CV of the individual C , values to be higher.…”

Section: Plasma Membrane Capacitymentioning

confidence: 99%

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Electropermeabilization and fluorescent tracer exchange: The role of whole‐cell capacitance

et al. 1995

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Transmembrane crossing of charged fluorescent tracers such as propidium iodide (PI) and carboxyfluorescein (CF) can be used to quantitate membrane permeabilization. Murine myeloma Sp2/0-Ag14 cells were loaded with CF (0.1 fmolkell) before electropulsation (0.5-3.0 kV/cm, 40 ps) in medium containing 25-50 pg/ml PI at 21-23°C. Cytograms of PI vs. CF fluorescence showed three readily distinguishable subpopulations: 1) intact living cells with CF but without PI (these form >95% of the prepulsed population), 2) transiently electropermeabilized but resealed cells showing both CF and low-level PI fluorescence, and 3) permanently permeabilized cells without CF but with very high PI fluorescence. Despite the ready influx of PI, the efflux of CF from transiently permeabilized cells was negligible and was insensitive to pulse parameters; however, electrically killed cells (subpopulation 3) lost all CF fluorescence and probably lost their cytoplasm. This difference in transmembrane passage of the dyes is best explained by binding of intracellular CF to macromolecules (and/or organelles). In isotonic "pulse medium," the membranes resealed after electropulsing with a time constant (73 of about 2 min. In 150 mOsm medium, resealing was faster (typically T~ -0.5 min). The population distribution of PI uptake [coefficient of variation (CV) > 40%] was very broad and could not be accounted for by the radius de-10%). The variability in PI uptake could be explained if the electrical energy of the charged membrane, which depends on the whole-cell capacitance (C,), was taken into account. Evaluation of the C, values with single-cell resolution was based on measurement of the electrical charging time constant of the plasma membrane by electrorotation. Key terms: Plasma membrane, electric breakdown, cytoplasm, fluorescent staining, propidium iodide, dye uptake, electrorotation, carboxyfluorescein, flow cytometry, cell viability Electropermeabilization allows the introduction of foreign substances (e.g., drugs, hormones, proteins, plasmids) into living cells (for reviews, see 27,44,45). This technique has been reported to be the most reliable for studying the effects of normally impermeant molecules on the regulation of cell metabolism. Other permeabilization methods (e.g., the use of bacterial toxins, detergents, hypertonic treatment) can markedly affect cell viability and cause undesirable leakage of the cytoplasm or extensive cell lysis ( 2 5 3 0 ) .Plasma membrane electropermeabilization within statistically representative cell samples can be assessed by measuring membrane crossing of tracer molecules in a flow cytometer. Extremely heterogeneous responses to electropulsing have been found in rather homogeneous (origin, size, morphology) cell samples despite the use of uniform electric fields (parallel plate electrodes) for several cell types (6,9,10,24). Flow cytometry of human red blood cells after electropulsation in the presence of FITCdextran (70 kDa) gave rise to at least three distinct subpopulations: ghosts with negligible upta...

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Section: Plasma Membrane Capacitymentioning

confidence: 99%

“…The 570 nm absorbance of the samples (including light scattering) was maintained at about 0.1. The time course of red fluorescence was followed continuously (resolution 0.1 s) from 1 min before to [10][11][12][13][14][15] min after field application (recording mode of the spectrophotometer).…”

Section: Electropermeabilization Of the Plasma Membranementioning

confidence: 99%

Electropermeabilization and fluorescent tracer exchange: The role of whole‐cell capacitance

et al. 1995

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“…The principle of measuring biomass by dielectric spectroscopy in terms of capacitance is that, at radio frequencies, the quantity of biomass is expressed as the contribution of intact and non-conducting cell membrane enclosures in suspension, within and beyond which there exist high concentrations of electrically conductive substances (Harris et al, 1987). Although the exploration and exploitation of the capacitance probe for online microbial cultivation has met with much success (Markx and Kell, 1995), research using this technique online during animal cell cultivation is still in its infancy (Davey et al, 1997), relying on valuable information from off-line tests (Cerckel et al, 1993;Davey et al, 1988;Mishima et al, 1991).…”

Section: Theoreticalmentioning

confidence: 99%

Specific heat flow rate: An on-line monitor and potential control variable of specific metabolic rate in animal cell culture that combines microcalorimetry with dielectric spectroscopy

Guan

Evans

Кемп

1998

Biotechnol. Bioeng.

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“…Dielectric spectroscopy at radio frequencies has already proven to be a useful tool for the estimation of the biomass levels of many different kinds of organisms (Harris et al, 1987;Davey et al, 1988Davey et al, , 1991Boulton et al, 1989;Kell & Todd, 1989;Stoicheva et al, 1989;Ferris et a f . , 1990;Kell et al, 1990;Peiialoza et al, 1991;Salter et al, 1990).…”

Section: Aberystwythmentioning

confidence: 99%

The permittistat: a novel type of turbidostat

Markx

Davey

Kell

1991

Journal of General Microbiology

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Baker's yeast was grown in a novel type of turbidostat in which the steady-state biomass level was controlled not by the optical turbidity but by the dielectric permittivity of the suspension at appropriate radio frequencies. Dry weight, fresh weight, the optical density at 600 nm, percentage viability (from methylene blue staining), bud count and ethanol concentration were measured off-line and the cell size distribution was recorded using flow cytometry. Any changes in the physiological properties of the yeast had a negligible effect on the ratio between the permittivity set (and measured) and the steady-state dry weight, fresh weight or optical density of the cultures. The permittistat was found to provide an extremely convenient means for carrying out turbidostatic culture.

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On the audio- and radio-frequency dielectric behaviour of anchorage-independent, mouse L929-derived LS fibroblasts

Cited by 46 publications

References 42 publications

Electropermeabilization and fluorescent tracer exchange: The role of whole‐cell capacitance

Electropermeabilization and fluorescent tracer exchange: The role of whole‐cell capacitance

Specific heat flow rate: An on-line monitor and potential control variable of specific metabolic rate in animal cell culture that combines microcalorimetry with dielectric spectroscopy

The permittistat: a novel type of turbidostat

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