Dielectric imaging for differentiation between cancer and inflammation in vivo.

In this study, we develop an in vivo dielectric imaging technique that measures capacitance using pin-type electrode arrays. Compared to normal tissues, cancer tissues exhibit higher capacitance values, allowing us to image the cancer region and monitor the chemotherapeutic effects of cancer in real-time. A comparison with the histopathological results shows that the in vivo dielectric imaging technique is able to detect small tumors (<3 mm) and tumor-associated changes. In addition, we demonstrate that cancer and inflammation may be distinguished by measuring the capacitance images at different frequencies. In contrast, the positron emission tomography using 2-[18F]-fluoro-2-deoxy-D-glucose was not capable of discriminating between cancer and inflammation.

Real-time and label-free monitoring of nanoparticle cellular uptake using capacitance-based assays.

Nanoparticles have shown great potential as vehicles for the delivery of drugs, nucleic acids, and therapeutic proteins; an efficient, high-throughput screening method to analyze nanoparticle interaction with the cytomembrane would substantially improve the efficiency and accuracy of the delivery. Here, we developed a capacitance sensor array that monitored the capacitance values of nanoparticle-treated cells in a real-time manner, without the need for labeling. Upon cellular uptake of the nanoparticles, a capacitance peak was observed at a low frequency (e.g., 100 Hz) as a function of time based on zeta potential changes. In the high frequency region (e.g., 15-20 kHz), the rate of decreasing capacitance slowed as a function of time compared to the cell growth control group, due to increased cytoplasm resistance and decreased membrane capacitance and resistance. The information provided by our capacitance sensor array will be a powerful tool for scientists designing nanoparticles for specific purposes.

Real-time monitoring of 3D cell culture using a 3D capacitance biosensor.

Three-dimensional (3D) cell cultures have recently received attention because they represent a more physiologically relevant environment compared to conventional two-dimensional (2D) cell cultures. However, 2D-based imaging techniques or cell sensors are insufficient for real-time monitoring of cellular behavior in 3D cell culture. Here, we report investigations conducted with a 3D capacitance cell sensor consisting of vertically aligned pairs of electrodes. When GFP-expressing human breast cancer cells (GFP-MCF-7) encapsulated in alginate hydrogel were cultured in a 3D cell culture system, cellular activities, such as cell proliferation and apoptosis at different heights, could be monitored non-invasively and in real-time by measuring the change in capacitance with the 3D capacitance sensor. Moreover, we were able to monitor cell migration of human mesenchymal stem cells (hMSCs) with our 3D capacitance sensor.

Real-time estimation of paracellular permeability of cerebral endothelial cells by capacitance sensor array.

Vascular integrity is important in maintaining homeostasis of brain microenvironments. In various brain diseases including Alzheimer's disease, stroke, and multiple sclerosis, increased paracellular permeability due to breakdown of blood-brain barrier is linked with initiation and progression of pathological conditions. We developed a capacitance sensor array to monitor dielectric responses of cerebral endothelial cell monolayer, which could be utilized to evaluate the integrity of brain microvasculature. Our system measured real-time capacitance values which demonstrated frequency- and time-dependent variations. With the measurement of capacitance at the frequency of 100 Hz, we could differentiate the effects of vascular endothelial growth factor (VEGF), a representative permeability-inducing factor, on endothelial cells and quantitatively analyse the normalized values. Interestingly, we showed differential capacitance values according to the status of endothelial cell monolayer, confluent or sparse, evidencing that the integrity of monolayer was associated with capacitance values. Another notable feature was that we could evaluate the expression of molecules in samples in our system with the reference of real-time capacitance values. We suggest that this dielectric spectroscopy system could be successfully implanted as a novel in vitro assay in the investigation of the roles of paracellular permeability in various brain diseases.

Real-time discrimination between proliferation and neuronal and astroglial differentiation of human neural stem cells.

Neural stem cells (NSCs) are characterized by a capacity for self-renewal, differentiation into multiple neural lineages, all of which are considered to be promising components for neural regeneration. However, for cell-replacement therapies, it is essential to monitor the process of in vitro NSC differentiation and identify differentiated cell phenotypes. We report a real-time and label-free method that uses a capacitance sensor array to monitor the differentiation of human fetal brain-derived NSCs (hNSCs) and to identify the fates of differentiated cells. When hNSCs were placed under proliferation or differentiation conditions in five media, proliferating and differentiating hNSCs exhibited different frequency and time dependences of capacitance, indicating that the proliferation and differentiation status of hNSCs may be discriminated in real-time using our capacitance sensor. In addition, comparison between real-time capacitance and time-lapse optical images revealed that neuronal and astroglial differentiation of hNSCs may be identified in real-time without cell labeling.

Real-time monitoring of adipocyte differentiation using a capacitance sensor array.

As obesity and its associated metabolic diseases become a worldwide epidemic, the demand for novel anti-obesity agents is increasing. We report a label-free and real-time monitoring method that uses a capacitance sensor array to screen anti-obesity agents. The results for the real-time capacitance of 3T3-L1 cells treated with 12 different chemicals extracted from natural products were consistent with the biochemical indicators of adipogenesis such as the expression of perilipin, the major protein coating the surface of lipid droplets in adipocytes. The data demonstrate that a capacitance change during adipocyte differentiation is closely associated with lipid accumulation in the cells, suggesting that adipocyte differentiation can be monitored in real time. This capacitance sensor might be used for label-free and real-time monitoring of adipocyte differentiation, and may facilitate the development of high throughput screening methods for anti-obesity drugs.

Capacitance-based assay for real-time monitoring of endocytosis and cell viability.

Label-free cell-based assays have emerged as a promising means for high-throughput screening. Most label-free sensors are based on impedance measurements that reflect the passive electrical properties of cells. Here we introduce a capacitance-based assay that measures the dielectric constant (capacitance) of biological cells, and demonstrate the feasibility of analyzing endocytosis and screening chemotherapeutic agents with this assay. Endocytosis induces a change in the zeta potential, leading to a change in the dielectric constant which enables real-time endocytosis monitoring using the capacitance sensor. Additionally, since the dielectric constant is proportional to cell radius and cell volume, cell viability can be estimated from the change in capacitance. Therefore, the capacitance sensor array can also be used for cytotoxicity testing for large-scale chemotherapeutic screening.

Capacitance-based real time monitoring of receptor-mediated endocytosis.

Receptor-mediated endocytosis is essential for targeted gene/drug delivery to a specific cell type. In this study, we developed a capacitance sensor to monitor receptor-mediated endocytosis in real time. The capacitance sensor was able to detect a capacitance peak in different cell lines during the internalization of adenoviruses or antibodies via receptor-mediated endocytosis. In contrast, the capacitance declined without a capacitance peak when nanoparticles were taken up via non-specific pinocytosis. Thus, our capacitance sensor represents a potential capacitance-based means of discrimination between receptor-mediated endocytosis and non-specific pinocytosis. Moreover, we developed a capacitance sensor array to demonstrate capacitance-based high-throughput screening. We showed that the capacitance sensor array could rapidly identify antibodies or ligands with high specificity for target molecules. We propose that the capacitance sensor array will provide a valuable tool for high-throughput screening.