Nitrogen-doped activated carbon composite electrode for deionization of phosphate removal and DFT model adsorption of phosphates.

Phosphate discharge in sewage can result in water eutrophication, posing a threat to aquatic ecosystems. Membrane capacitive deionization (MCDI) has demonstrated outstanding performance and significant potential for salt removal and nutrient recovery. In this study, a nitrogen-doped activated carbon electrode material (NAC) was synthesized through one-step pyrolysis to selectively remove phosphate from MCDI. At a voltage of 1.2V, a flow rate of 20 mL/min, and a pH of 6.51, the phosphate adsorption capacity of the NAC electrode was determined to be 1.60 mg/g. The study revealed that NAC pHpzc increased from 4.14 to 6.44, effectively broadening the pH range for phosphate removal. In the presence of competing ions (NO3-, Cl-, and SO42-) at a concentration of 0.5 M, the electroadsorption capacity of phosphate decreased to 1.21 mg/g, 1.14 mg/g, and 1.02 mg/g, respectively. The kinetic parameters of adsorption indicated that NAC electroadsorbed phosphate through physical adsorption, with the maximum adsorption capacity achieved at 303K. Data from the Freundlich isothermal model suggested that phosphate adsorption by the NAC electrode involves a multilayer adsorption process. A carbon structure model of density functional theory (DFT), incorporating doped nitrogen, was constructed based on XPS analysis. Following nitrogen doping, the electrostatic potential (ESP) of unsaturated carbon atoms became more positive, enhancing the ability of nitrogen-doped activated carbon to adsorb phosphate. This study provides compelling evidence that nitrogen doping facilitates the adsorption of phosphate by carbon materials.

Effect of colloidal magnetite (Fe3O4) nanoparticles on the electrical characteristics of the azolectin bilayer in a static inhomogeneous magnetic field.

This work is devoted to the study of the combined effects of applied magnetic field and MNPs on the electrical characteristics of bilayer lipid membranes. We present results of the study of electrical parameters of azolectin membranes in a static inhomogeneous magnetic field at the one-sided addition of positively charged quasi-spherical superparamagnetic magnetite nanoparticles with a diameter of about 4 nm. The magnet was located at different distances from the membrane, and the magnetic field attracted the nanoparticles to the membrane surface with different strengths. We observed three pronounced effects that depended on the external magnetic field. Firstly, after addition of nanoparticles in a magnetic field, the conductance of the membranes increased. A smooth increase in conductance was accompanied in some cases by the appearance of current jumps, which can be associated with the formation of through pores with a radius of no more than 1 nm. The conductance increased with increasing magnetic field gradient. Secondly, at zero command voltage, a negative current through the membrane was observed. Although our experiments did not allow us to unambiguously determine which particles create this current, we believe that this current is associated with the penetration of particles through the membrane. This effect intensified with increasing magnetic field gradient. Thirdly, we observed a sharp change in the nonlinear dependence of capacitance on voltage associated both with the change in the surface potential of the azolectin membrane and with the effect of MNP binding to the membrane surface on the apparent membrane capacitance.

Prazosin Potentiates Mast Cell-Stabilizing Property of Adrenaline.

BACKGROUND/AIMS: Adrenaline quickly inhibits the release of histamine from mast cells. Besides ß2-adrenergic receptors, several in vitro studies also indicate the involvement of α-adrenergic receptors in the process of exocytosis. Since exocytosis in mast cells can be detected electrophysiologically by the changes in the membrane capacitance (Cm), its continuous monitoring in the presence of drugs would determine their mast cell-stabilizing properties. METHODS: Employing the whole-cell patch-clamp technique in rat peritoneal mast cells, we examined the effects of adrenaline on the degranulation of mast cells and the increase in the Cm during exocytosis. We also examined the degranulation of mast cells in the presence or absence of α-adrenergic receptor agonists or antagonists. RESULTS: Adrenaline dose-dependently suppressed the GTP-γ-S-induced increase in the Cm and inhibited the degranulation from mast cells, which was almost completely erased in the presence of butoxamine, a ß2-adrenergic receptor antagonist. Among α-adrenergic receptor agonists or antagonists, high dose prazosin, a selective α1-adrenergic receptor antagonist, significantly reduced the ratio of degranulating mast cells and suppressed the increase in the Cm. Additionally, prazosin augmented the inhibitory effects of adrenaline on the degranulation of mast cells. CONCLUSION: This study provided electrophysiological evidence for the first time that adrenaline dose-dependently inhibited the process of exocytosis, confirming its usefulness as a potent mast cell-stabilizer. The pharmacological blockade of α1-adrenergic receptor by prazosin synergistically potentiated such mast cell-stabilizing property of adrenaline, which is primarily mediated by ß2-adrenergic receptors.

Relationship between ion currents and membrane capacitance in canine ventricular myocytes.

Current density, the membrane current value divided by membrane capacitance (Cm), is widely used in cellular electrophysiology. Comparing current densities obtained in different cell populations assume that Cm and ion current magnitudes are linearly related, however data is scarce about this in cardiomyocytes. Therefore, we statistically analyzed the distributions, and the relationship between parameters of canine cardiac ion currents and Cm, and tested if dividing original parameters with Cm had any effect. Under conventional voltage clamp conditions, correlations were high for IK1, moderate for IKr and ICa,L, while negligible for IKs. Correlation between Ito1 peak amplitude and Cm was negligible when analyzing all cells together, however, the analysis showed high correlations when cells of subepicardial, subendocardial or midmyocardial origin were analyzed separately. In action potential voltage clamp experiments IK1, IKr and ICa,L parameters showed high correlations with Cm. For INCX, INa,late and IKs there were low-to-moderate correlations between Cm and these current parameters. Dividing the original current parameters with Cm reduced both the coefficient of variation, and the deviation from normal distribution. The level of correlation between ion currents and Cm varies depending on the ion current studied. This must be considered when evaluating ion current densities in cardiac cells.

Electrical signature of heterogeneous human mesenchymal stem cells.

Human mesenchymal stem cells (hMSCs) have gained traction in transplantation therapy due to their immunomodulatory, paracrine, immune-evasive, and multipotent differentiation potential. The inherent heterogeneity of hMSCs poses a challenge for therapeutic treatments and necessitates the identification of robust biomarkers to ensure reproducibility in both in vivo and in vitro experiments. In this study, we utilized dielectrophoresis (DEP), a label-free electrokinetic phenomenon, to investigate the heterogeneity of hMSCs derived from bone marrow (BM) and adipose tissue (AD). The electrical properties of BM-hMSCs were compared to homogeneous mouse fibroblasts (NIH-3T3), human fibroblasts (WS1), and human embryonic kidney cells (HEK-293). The DEP profile of BM-hMSCs differed most from HEK-293 cells. We compared the DEP profiles of BM-hMSCs and AD-hMSCs and found that they have similar membrane capacitances, differing cytoplasm conductivity, and transient slopes. Inducing both populations to differentiate into adipocyte and osteoblast cells revealed that they behave differently in response to differentiation-inducing cytokines. Histology and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analyses of the differentiation-related genes revealed differences in heterogeneity between BM-hMSCs and AD-hMSCs. The differentiation profiles correlate well with the DEP profiles developed and indicate differences in the heterogeneity of BM-hMSCs and AD-hMSCs. Our results demonstrate that using DEP, membrane capacitance, cytoplasm conductivity, and transient slope can uniquely characterize the inherent heterogeneity of hMSCs to guide robust and reproducible stem cell transplantation therapies.

Photomanipulation of Minimal Synthetic Cells: Area Increase, Softening, and Interleaflet Coupling of Membrane Models Doped with Azobenzene-Lipid Photoswitches.

Light can effectively interrogate biological systems in a reversible and physiologically compatible manner with high spatiotemporal precision. Understanding the biophysics of photo-induced processes in bio-systems is crucial for achieving relevant clinical applications. Employing membranes doped with the photolipid azobenzene-phosphatidylcholine (azo-PC), a holistic picture of light-triggered changes in membrane kinetics, morphology, and material properties obtained from correlative studies on cell-sized vesicles, Langmuir monolayers, supported lipid bilayers, and molecular dynamics simulations is provided. Light-induced membrane area increases as high as ≈25% and a ten-fold decrease in the membrane bending rigidity is observed upon trans-to-cis azo-PC isomerization associated with membrane leaflet coupling and molecular curvature changes. Vesicle electrodeformation measurements and atomic force microscopy reveal that trans azo-PC bilayers are thicker than palmitoyl-oleoyl phosphatidylcholine (POPC) bilayers but have higher specific membrane capacitance and dielectric constant suggesting an increased ability to store electric charges across the membrane. Lastly, incubating POPC vesicles with azo-PC solutions results in the insertion of azo-PC in the membrane enabling them to become photoresponsive. All these results demonstrate that light can be used to finely manipulate the shape, mechanical and electric properties of photolipid-doped minimal cell models, and liposomal drug carriers, thus, presenting a promising therapeutic alternative for the repair of cellular disorders.

Electrostatic Potentials Caused by the Release of Protons from Photoactivated Compound Sodium 2-Methoxy-5-nitrophenyl Sulfate at the Surface of Bilayer Lipid Membrane.

Lateral transport and release of protons at the water-membrane interface play crucial roles in cell bioenergetics. Therefore, versatile techniques need to be developed for investigating as well as clarifying the main features of these processes at the molecular level. Here, we experimentally measured the kinetics of binding of protons released from the photoactivated compound sodium 2-methoxy-5-nitrophenyl sulfate (MNPS) at the surface of a bilayer lipid membrane (BLM). We developed a theoretical model of this process describing the damage of MNPS coupled with the release of the protons at the membrane surface, as well as the exchange of MNPS molecules and protons between the membrane and solution. We found that the total change in the boundary potential difference across the membrane, ∆ϕb, is the sum of opposing effects of adsorption of MNPS anions and release of protons at the membrane-water interface. Steady-state change in the ∆ϕb due to protons decreased with the concentration of the buffer and increased with the pH of the solution. The change in the concentration of protons evaluated from measurements of ∆ϕb was close to that in the unstirred water layer near the BLM. This result, as well as rate constants of the proton exchange between the membrane and the bulk solution, indicated that the rate-limiting step of the proton surface to bulk release is the change in the concentration of protons in the unstirred layer. This means that the protons released from MNPS remain in equilibrium between the BLM surface and an adjacent water layer.

High-accuracy gastric cancer cell viability evaluation based on multi-impedance spectrum characteristics.

The increasing attention to precision medicine is widely paid to greatly rise the cure rate of cancer. Improving the stability and accuracy of cancer cell viability evaluation is one of the keys for precision medicine, as excess dosage of anti-cancer drugs not only kills the cancer cells, but also does harm to normal cells. Electrochemical impedance sensing (EIS) method is well known as a label-free, non-invasive approach for real-time, online monitoring of cell viability. However, the existing EIS methods using single-frequency impedances cannot reflect the comprehensive information of cellular impedance spectroscopy (CIS), ultimately leading to a poor stability and low accuracy of cancer cell viability evaluation. In this paper, we proposed a multi-frequency approach for improving the stability and accuracy of cancer cell viability evaluation based on multi-physical properties of CIS, including cell adhesion state and cell membrane capacitance. The results show that the mean relative error of multi-frequency method is reduced by 50% compared with single-frequency method, while the maximum relative error of the former is 7∼fold smaller than that of the latter. The accuracy of cancer cell viability evaluation is up to 99.6%.

Comprehensive multiparameter evaluation of platelet function using a highly sensitive membrane capacitance sensor.

An accurate and comprehensive assessment of platelet function is essential for managing patients who receive antiplatelet therapies or require platelet transfusion either for treating active bleeding or for prophylaxis. Platelets contribute to clotting by undergoing a series of highly regulated functional responses including adhesion, spreading, granular secretion, aggregation, and cytoskeletal contraction. However, current platelet function assays evaluate only partial aspects of this intricate process and often under non-physiological testing conditions. Herein, we describe the development of a new approach to measure multiple key platelet function-related parameters, in a more physiologically relevant ex vivo semi-rigid microenvironment using a membrane capacitance sensor (MCS). MCS response to clotting provided three sensing parameters with sensitivities towards platelet counts, stimulation strengths, and activation pathways. Live confocal fluorescent imaging of stimulated platelets on MCS suggests that the presented system can readily and accurately convert the dynamics of cytoskeletal reorganization into analyzable electrical signals. Together, this new completely electrical sensing platform can be a promising diagnostic venue to recognize the impairment of primary hemostatic functions, evaluate the efficacy of therapeutic interventions, and gain further insights into the mechanisms of platelets in hemostasis and thrombosis.

Changes in Electrical Capacitance of Cell Membrane Reflect Drug Partitioning-Induced Alterations in Lipid Bilayer.

The plasma membrane is a lipid bilayer that establishes the outer boundary of a living cell. The composition of the lipid bilayer influences the membrane's biophysical properties, including fluidity, thickness, permeability, phase behavior, charge, elasticity, and formation of flat sheet or curved structures. Changes in the biophysical properties of the membrane can be occasioned when new entities, such as drug molecules, are partitioned in the bilayer. Therefore, assessing drugs for their effect on the biophysical properties of the lipid bilayer of a cell membrane is critical to understanding specific and non-specific drug action. Previously, we reported a non-invasive technique for real-time characterization of cellular dielectric properties, such as membrane capacitance and cytoplasmic conductivity. In this study, we discuss the potential application of the technique in assessing the biophysical properties of the cell membrane in response to interaction with amiodarone compared to aspirin/acetylsalicylic acid and glucose. Amiodarone is a potent drug used to treat cardiac arrhythmia, but it also exerts various non-specific effects. Compared to aspirin and glucose, we measured a rapid and higher magnitude increase in membrane capacitance on cells under amiodarone treatment. Increased membrane capacitance induced by aspirin and glucose quickly returned to baseline in 15 s, while amiodarone-induced increased capacitance sustained and decreased slowly, approaching baseline or another asymptotic limit in ~2.5 h. Because amiodarone has a strong lipid partitioning property, we reason that drug partitioning alters the lipid bilayer context and subsequently reduces bilayer thickness, leading to an increase in the electrical capacitance of the cell membrane. The presented microfluidic system promises a new approach to assess drug-membrane interactions and delineate specific and non-specific actions of the drug on cells.

Liposome Deformation Induced by Membrane-Binding Peptides.

This paper presents an investigation of liposome deformation and shape distortion using four membrane-binding peptides: TAT and C105Y as cell-penetrating peptides (CPPs), and melittin and ovispirin as antimicrobial peptides (AMPs). Liposome deformation was monitored utilizing fluorescent microscopy, while the binding of peptides to the DOPC membrane was estimated through capacitance measurements. The degree of liposome deformation and shape distortion was found to be higher for the CPPs compared to the AMPs. Additionally, it was observed that C105Y did not induce liposome rupture, unlike the other three peptides. We propose that these variations in liposome distortion may be attributed to differences in secondary structure, specifically the presence of an α-helix or random coil. Our studies offer insight into the use of peptides to elicit control of liposome architecture and may offer a promising approach for regulating the bodies of liposomal molecular robots.

A new approach to the determination of tubular membrane capacitance: passive membrane electrical properties under reduced electrical conductivity of the extracellular solution.

The transverse-axial tubular system (tubular system) of cardiomyocytes plays a key role in excitation-contraction coupling. To determine the area of the tubular membrane in relation to the area of the surface membrane, indirect measurements through the determination of membrane capacitances are currently used in addition to microscopic methods. Unlike existing electrophysiological methods based on an irreversible procedure (osmotic shock), the proposed new approach uses a reversible short-term intermittent increase in the electrical resistance of the extracellular medium. The resulting increase in the lumen resistance of the tubular system makes it possible to determine separate capacitances of the tubular and surface membranes. Based on the analysis of the time course of the capacitive current, computational relations were derived to quantify the elements of the electrical equivalent circuit of the measured cardiomyocyte including both capacitances. The exposition to isotonic low-conductivity sucrose solution is reversible which is the main advantage of the proposed approach allowing repetitive measurements on the same cell under control and sucrose solutions. Experiments on rat ventricular cardiomyocytes (n = 20) resulted in the surface and tubular capacitance values implying the fraction of tubular capacitance/area of 0.327 ± 0.018. We conclude that the newly proposed method provides results comparable to the data obtained by the currently used detubulation method and, in addition, by being reversible, allows repeated evaluation of surface and tubular membrane parameters on the same cell.

Response of a neuronal network computational model to infrared neural stimulation.

Infrared neural stimulation (INS), as a novel form of neuromodulation, allows modulating the activity of nerve cells through thermally induced capacitive currents and thermal sensitivity ion channels. However, fundamental questions remain about the exact mechanism of INS and how the photothermal effect influences the neural response. Computational neural modeling can provide a powerful methodology for understanding the law of action of INS. We developed a temperature-dependent model of ion channels and membrane capacitance based on the photothermal effect to quantify the effect of INS on the direct response of individual neurons and neuronal networks. The neurons were connected through excitatory and inhibitory synapses and constituted a complex neuronal network model. Our results showed that a slight increase in temperature promoted the neuronal spikes and enhanced network activity, whereas the ultra-temperature inhibited neuronal activity. This biophysically based simulation illustrated the optical dose-dependent biphasic cell response with capacitive current as the core change condition. The computational model provided a new sight to elucidate mechanisms and inform parameter selection of INS.

Membrane capacitance and characteristic frequency are associated with contractile properties of skeletal muscle.

The cell membrane capacitance (Cm) and characteristic frequencies (fc) of tissues can be obtained using segmental bioelectrical impedance spectroscopy (S-BIS). Higher Cm and lower fc are associated with a larger surface area of skeletal muscle fibers with T-tubules in the tissues. Muscle fiber membrane is one of the major physiological factors that influence surface electromyograms (EMGs) as well as the number of recruited motor units so that the amplitude of surface EMG may be correlated with Cm and fc. The aim of the current study was to examine the association of fc or Cm in the lower leg with contractile and neuromuscular properties in the plantar flexors. We analyzed data from 59 participants (29 women) aged 21-83 years. The Cm, fc, and intracellular water (ICW) in the lower leg were obtained using S-BIS. We measured electrical-evoked torque, maximal voluntary contraction (MVC) torque, and amplitude of EMG normalized by the M wave during MVC contraction. The high Cm group had a significantly lower fc and significantly higher MVC torque, estimated maximum torque, twitch torque, and root mean square (RMS) of EMG normalized by the M wave (EMG:M) in the musculus triceps surae compared to the low Cm group (P < 0.05). Cm was positively and fc was negatively correlated with the nRMS of EMG:M in the triceps surae (P < 0.05). S-BIS recordings can be used to detect changes in skeletal muscle membrane capacitance, which may provide insights into the number of T-tubules. The muscle capacitance measured with S-BIS can be predictive of muscle force generation.

Pyridoxine Synergistically Potentiates Mast Cell-Stabilizing Property of Ascorbic Acid.

BACKGROUND/AIMS: Besides their physiological properties, vitamins, such as vitamin C (ascorbic acid) and B6 (pyridoxine), ameliorate the symptoms of allergic disorders. Because exocytosis in mast cells can be detected electrophysiologically by the changes in the membrane capacitance (Cm), its continuous monitoring in the presence of these vitamins would determine their mast cell-stabilizing, anti-allergic properties. METHODS: Employing the whole-cell patch-clamp technique in rat peritoneal mast cells, we examined the effects of ascorbic acid and pyridoxine on the degranulation of mast cells and the increase in the Cm during exocytosis. RESULTS: Both ascorbic acid and pyridoxine dose-dependently suppressed the GTP-γ-S-induced increase in the Cm and inhibited the degranulation from mast cells. Surprisingly enough, relatively low concentrations of pyridoxine (1, 2 mM) synergistically enhanced the suppressive effect of 2 mM ascorbic acid on mast cell degranulation. CONCLUSION: These results provided electrophysiological evidence for the first time that ascorbic acid and pyridoxine inhibited the process of exocytosis in a dose-dependent manner. At relatively lower concentrations, these vitamins were not enough to stabilize mast cells. However, such concentrations of pyridoxine synergistically potentiated the mast cell-stabilizing property of ascorbic acid.

Optocapacitance: physical basis and its application.

The observation that membrane capacitance increases with temperature has led to the development of new methods of neuronal stimulation using light. The optocapacitive effect refers to a light-induced change in capacitance produced by the heating of the membrane through a photothermal effect. This change in capacitance manifests as a current, named optocapacitive current that depolarizes cells and therefore can be used to stimulate excitable tissues. Here, we discuss how optocapacitance arises from basic membrane properties, the characteristics of the optocapacitive current, its use for neuronal stimulation, and the challenges for its application in vivo.

Characterization of Extra-Cellular Vesicle Dielectrophoresis and Estimation of Its Electric Properties.

Dielectrophoresis (DEP) refers to a type of electrical motion of dielectric particles. Because DEP is caused by particle polarization, it has been utilized to characterize particles. This study investigated the DEP of three types of exosomes, namely bovine milk, human breast milk, and human breast cancer exosomes. Exosomes are kinds of extracellular vesicles. The crossover frequencies of the exosomes were determined by direct observation of their DEPs. Consequently, bovine and human milk exosomes showed similar DEP properties, whereas the cancer exosomes were significantly different from the others. The membrane capacitance and conductivity of the exosomes were estimated using determined values. A significant difference was observed between bovine and human milk exosomes on their membrane capacitance. It was revealed that the membrane capacitances of human breast milk and human breast cancer exosomes were almost identical to those of their host cells and the conductivity of the exosomes were much lower than that of the host cell. Based on these results, DEP separation of the human breast milk and cancer exosomes was demonstrated. These results imply that DEP can be utilized to separate and identify cancer exosomes rapidly. Additionally, our method can be utilized to estimate the electric property of other types of extracellular vesicles.

Fraction of the T-Tubular Membrane as an Important Parameter in Cardiac Cellular Electrophysiology: A New Way of Estimation.

The transverse-axial tubular system (t-tubules) plays an essential role in excitation-contraction coupling in cardiomyocytes. Its remodelling is associated with various cardiac diseases. Numerous attempts were made to analyse characteristics essential for proper understanding of the t-tubules and their impact on cardiac cell function in health and disease. The currently available methodical approaches related to the fraction of the t-tubular membrane area produce diverse data. The widely used detubulation techniques cause irreversible cell impairment, thus, distinct cell samples have to be used for estimation of t-tubular parameters in untreated and detubulated cells. Our proposed alternative method is reversible and allows repetitive estimation of the fraction of t-tubular membrane (f t) in cardiomyocytes using short-term perfusion of the measured cell with a low-conductive isotonic sucrose solution. It results in a substantial increase in the electrical resistance of t-tubular lumen, thus, electrically separating the surface and t-tubular membranes. Using the whole-cell patch-clamp measurement and the new approach in enzymatically isolated rat atrial and ventricular myocytes, a set of data was measured and evaluated. The analysis of the electrical equivalent circuit resulted in the establishment of criteria for excluding measurements in which perfusion with a low conductivity solution did not affect the entire cell surface. As expected, the final average f t in ventricular myocytes (0.337 ± 0.017) was significantly higher than that in atrial myocytes (0.144 ± 0.015). The parameter f t could be estimated repetitively in a particular cell (0.345 ± 0.021 and 0.347 ± 0.023 in ventricular myocytes during the first and second sucrose perfusion, respectively). The new method is fast, simple, and leaves the measured cell intact. It can be applied in the course of experiments for which it is useful to estimate both the surface and t-tubular capacitance/area in a particular cell.

A dynamic clamp protocol to artificially modify cell capacitance.

Dynamics of excitable cells and networks depend on the membrane time constant, set by membrane resistance and capacitance. Whereas pharmacological and genetic manipulations of ionic conductances of excitable membranes are routine in electrophysiology, experimental control over capacitance remains a challenge. Here, we present capacitance clamp, an approach that allows electrophysiologists to mimic a modified capacitance in biological neurons via an unconventional application of the dynamic clamp technique. We first demonstrate the feasibility to quantitatively modulate capacitance in a mathematical neuron model and then confirm the functionality of capacitance clamp in in vitro experiments in granule cells of rodent dentate gyrus with up to threefold virtual capacitance changes. Clamping of capacitance thus constitutes a novel technique to probe and decipher mechanisms of neuronal signaling in ways that were so far inaccessible to experimental electrophysiology.

Impedance-based non-invasive assay for ocular damage prediction on in vitro 3D reconstructed human corneal epithelium.

Reconstructed human cornea-like epithelium (RhCE) holds unprecedented promise for toxicological analyses and the replacement of animal use. However, current standards to evaluate potential ocular irritancy present a major downfall, the need to invasively alter tissue samples to evaluate cell viability. In this study, the applicability of impedance analysis was validated by monitoring the change in cell capacitance during tissue maturation and before and after chemical application using coupled electrodes. Our results indicate that cell maturation on RhCE models can be evaluated during model production using capacitance sensing offering a faster and simpler quality control criteria for RhCE model usability. Additionally, cell capacitance resulted to be more sensitive in detecting slight cell damages than methods based on cell metabolism, and when integrated into OECD-approved testing strategies, capacitance sensing performed as good as currently accepted methodologies displaying 66% sensitivity, 100% specificity and 83% accuracy when evaluated at 300 Hz. In summary, a quantitative analysis to predict in vivo ocular irritation based on changes in RhCE capacitance by impedance spectroscopy is suggested. This methodology represents a non-invasive and non-destructive alternative that would enable the monitoring of reversible effects or repeated dose toxicity.