Micro hole-based cell chip with impedance spectroscopy

Electric fields can be used for the characterisation and manipulation of single biological cells. One approach to avoid the effect of electrode polarisation is to position cells on micro holes and to apply the electrical fields via the micro holes. For a correct characterisation and optimal manipulation, the electrical properties of the micro hole/cell interface must be understood. In this article, the electrical characteristics of a micro hole-based cell chip were investigated. By FEM simulation, it was estimated that the impedance measurement with micro hole-based chip is most dependent on the cell adhesion/spread rather than the intra-cellular space (contribution of intra-cellular space to the total impedance: 0.07% at 1 kHz, 0.3% at 1 MHz). The effective frequency range in which the impedance related with cell state on the hole considerably influences total measured impedance was below several kiloHertz. From the experiments, it was shown that the impedance of cell cultured on the hole at the low frequency range is increased during the increase of cultivation period, but is sensitively decreased after applying only several nanolitres of culture medium including 5% dimethlysulfoxide. This micro hole-based chip has a potential for monitoring the cell growth and the membrane integrity of even single cell without any labelling.     

Origin of changes in electrical impedance during the growth and fermentation process of yeast in batch culture

The electrical impedance of the culture medium shows complex changes during the growth and fermentation process of yeast, and this prevents its possible application for the monitoring of certain yeast activities. Clarification of the mechanism of such changes is thus essential for practical use. As a first step toward this aim, the impedance, yeast concentration, and pH of a batch culture medium were measured using special cells with two compartments and also the usual type of cell with one compartment. In the special cells, the yeast was cultured in one compartment only. Conducting ions and nonconducting substances diffused through an intermediate porous membrane sandwiched between the two compartments. The impedances of the two compartments were measured simultaneously by the four-electrode method. The main mechanism responsible for increasing the impedance was the conducting ions produced by the yeast extract added as a nutrient to the culture broth by certain nonconducting substances during the process of growth. The increase in the yeast concentration was also a minor factor increasing the impedance. These increases surpassed the impedance decrease caused by the increase of H(+) ions produced by some accumulated acidic substances, and the impedance thus increased.     

Determination of Mammalian Cell Counts,Cell Size and Cell Health Using the Moxi Z Mini Automated Cell Counter

Particle and cell counting is used for a variety of applications including routine cell culture, hematological analysis, and industrial controls^1-5. A critical breakthrough in cell/particle counting technologies was the development of the Coulter technique by Wallace Coulter over 50 years ago. The technique involves the application of an electric field across a micron-sized aperture and hydrodynamically focusing single particles through the aperture. The resulting occlusion of the aperture by the particles yields a measurable change in electric impedance that can be directly and precisely correlated to cell size/volume. The recognition of the approach as the benchmark in cell/particle counting stems from the extraordinary precision and accuracy of its particle sizing and counts, particularly as compared to manual and imaging based technologies (accuracies on the order of 98% for Coulter counters versus 75-80% for manual and vision-based systems). This can be attributed to the fact that, unlike imaging-based approaches to cell counting, the Coulter Technique makes a true three-dimensional (3-D) measurement of cells/particles which dramatically reduces count interference from debris and clustering by calculating precise volumetric information about the cells/particles. Overall this provides a means for enumerating and sizing cells in a more accurate, less tedious, less time-consuming, and less subjective means than other counting techniques⁶.Despite the prominence of the Coulter technique in cell counting, its widespread use in routine biological studies has been prohibitive due to the cost and size of traditional instruments. Although a less expensive Coulter-based instrument has been produced, it has limitations as compared to its more expensive counterparts in the correction for "coincidence events" in which two or more cells pass through the aperture and are measured simultaneously. Another limitation with existing Coulter technologies is the lack of metrics on the overall health of cell samples. Consequently, additional techniques must often be used in conjunction with Coulter counting to assess cell viability. This extends experimental setup time and cost since the traditional methods of viability assessment require cell staining and/or use of expensive and cumbersome equipment such as a flow cytometer.The Moxi Z mini automated cell counter, described here, is an ultra-small benchtop instrument that combines the accuracy of the Coulter Principle with a thin-film sensor technology to enable precise sizing and counting of particles ranging from 3-25 microns, depending on the cell counting cassette used. The M type cassette can be used to count particles from with average diameters of 4 - 25 microns (dynamic range 2 - 34 microns), and the Type S cassette can be used to count particles with and average diameter of 3 - 20 microns (dynamic range 2 - 26 microns). Since the system uses a volumetric measurement method, the 4-25 microns corresponds to a cell volume range of 34 - 8,180 fL and the 3 - 20 microns corresponds to a cell volume range of 14 - 4200 fL, which is relevant when non-spherical particles are being measured. To perform mammalian cell counts using the Moxi Z, the cells to be counted are first diluted with ORFLO or similar diluent. A cell counting cassette is inserted into the instrument, and the sample is loaded into the port of the cassette. Thousands of cells are pulled, single-file through a "Cell Sensing Zone" (CSZ) in the thin-film membrane over 8-15 seconds. Following the run, the instrument uses proprietary curve-fitting in conjunction with a proprietary software algorithm to provide coincidence event correction along with an assessment of overall culture health by determining the ratio of the number of cells in the population of interest to the total number of particles. The total particle counts include shrunken and broken down dead cells, as well as other debris and contaminants. The results are presented in histogram format with an automatic curve fit, with gates that can be adjusted manually as needed.Ultimately, the Moxi Z enables counting with a precision and accuracy comparable to a Coulter Z2, the current gold standard, while providing additional culture health information. Furthermore it achieves these results in less time, with a smaller footprint, with significantly easier operation and maintenance, and at a fraction of the cost of comparable technologies.     

Embryonic Carcinoma Cells Show Specific Dielectric Resistance Profiles during Induced Differentiation

Induction of differentiation in cancer stem cells by drug treatment represents an important approach for cancer therapy. The understanding of the mechanisms that regulate such a forced exit from malignant pluripotency is fundamental to enhance our knowledge of tumour stability. Certain nucleoside analogues, such as 2′-deoxy-5-azacytidine and 1β-arabinofuranosylcytosine, can induce the differentiation of the embryonic cancer stem cell line NTERA 2 D1 (NT2). Such induced differentiation is associated with drug-dependent DNA-damage, cellular stress and the proteolytic depletion of stem cell factors. In order to further elucidate the mode of action of these nucleoside drugs, we monitored differentiation-specific changes of the dielectric properties of growing NT2 cultures using electric cell-substrate impedance sensing (ECIS). We measured resistance values of untreated and retinoic acid treated NT2 cells in real-time and compared their impedance profiles to those of cell populations triggered to differentiate with several established substances, including nucleoside drugs. Here we show that treatment with retinoic acid and differentiation-inducing drugs can trigger specific, concentration-dependent changes in dielectric resistance of NT2 cultures, which can be observed as early as 24 hours after treatment. Further, low concentrations of nucleoside drugs induce differentiation-dependent impedance values comparable to those obtained after retinoic acid treatment, whereas higher concentrations induce proliferation defects. Finally, we show that impedance profiles of substance-induced NT2 cells and those triggered to differentiate by depletion of the stem cell factor OCT4 are very similar, suggesting that reduction of OCT4 levels has a dominant function for differentiation induced by nucleoside drugs and retinoic acid. The data presented show that NT2 cells have specific dielectric properties, which allow the early identification of differentiating cultures and real-time label-free monitoring of differentiation processes. This work might provide a basis for further analyses of drug candidates for differentiation therapy of cancers.     

Cell placement and guidance on substrates for neurochip interfaces

Interface devices such as integrated planar patch‐clamp chips are being developed to study the electrophysiological activity of neuronal networks grown in vitro. The utility of such devices will be dependent upon the ability to align neurons with interface features on the chip by controlling neuronal placement and by guiding cell connectivity. In this paper, we present a strategy to accomplish this goal. Patterned chemical modification of SiN surfaces with poly‐d‐lysine transferred from PDMS stamps was used to promote adhesion and guidance of cryo‐preserved primary rat cortical neurons. We demonstrate that these neurons can be positioned and grown over microhole features which will ultimately serve as patch‐clamp interfaces on the chip. Biotechnol. Bioeng. 2010; 105: 368–373. © 2009 Wiley Periodicals, Inc.     

Synchronization of cell cycle of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> by using a cell chip platform

Cell synchrony is a critical requirement for the study of eukaryotic cells. Although several chemical and genetic methods of cell cycle synchronization are currently available, they have certain limitations, such as unnecessary perturbations to cells. We developed a novel cell cycle synchronization method that is based on a cell chip platform. The budding yeast, Saccharomyces cerevisiae, is a simple but useful model system to study cell biology and shares many similar features with higher eukaryotic cells. Single yeast cells were individually captured in the wells of a specially designed cell chip platform. When released from the cell chip, the yeast cells were synchronized, with all cells in the G1 phase. This method is non-invasive and causes minimal chemical and biological damage to cells. The capture and release of cells using cells chips with microwells of specific dimensions allows for the isolation of cells of a particular size and shape; this enables the isolation of cells of a given phase, because the size and shape of yeast cells vary with the phase of the cell cycle. To test the viability of synchronized cells, the yeast cells captured in the cell chip platform were assessed for response to mating pheromone (α-factor). The synchronized cells isolated using the cell chip were capable of mediating the mating signaling response and exhibited a dynamic and robust response behavior. By changing the dimensions of the well of the cell chip, cells of other cell cycle phases can also be isolated.     

植物微流芯片——一种实时定量监测生长发育的 高通量整合分析平台

生长发育是一个复杂的动态过程, 了解其发生细节是生命科学研究的重要内容。最新发展起来的微流芯片技术为实现这个目标提供了新的途径。动物及微生物中的应用表明, 该技术兼有实时定量监测和高通量整合处理的优势。在植物研究领域, 用针对根生长特点和要求设计的根微流芯片结合荧光共振能量转移探针已经成功地检测出拟南芥(Arabidopsis thaliana)根细胞内葡萄糖和游离的Ca²⁺、Zn²⁺的浓度。随着各种底物特异的荧光共振能量转移探针的开发和应用, 根微流芯片还可以用来检测植物细胞内激素或其它代谢中间产物的浓度及其动态变化过程。不仅如此, 以微流芯片为基础发展起来的Plant Chip和Tip Chip则为研究植物与微生物的相互作用以及植物花粉管极性生长和细胞分裂分化提供了理想的平台。作为了解遗传因素或环境刺激导致细胞生命活动变化细节的有力工具, 微流芯片技术有望为植物研究领域带来更多新的进展和突破。     

Patterning of embryonic stem cells using the bio flip chip

Cell-cell interactions consisting of diffusible signaling and cell-cell contact (juxtacrine signaling) are important in numerous biological processes such as tumor growth, stem cell differentiation, and stem cell self-renewal. A number of methods currently exist to modulate cell signaling in vitro. One method of modulating the total amount of diffusible signaling is to vary the cell seeding density during culture. Due to the random nature of cell seeding, this results in considerable variation in the actual cell-cell spacing and amount of cell-cell contact, and cannot prescribe the local environment. A more specific approach for modulating cell signaling is to use molecular inhibitors or genetic approaches to knock down specific signaling proteins, but both of these methods are best suited to manipulating small numbers of molecules. Here, we demonstrate a new approach to modulating cell-cell signaling that modulates the local environment of a cluster of cells by placing different numbers of cells at desired locations on a substrate. This method provides a complementary way to control the local diffusible and juxtacrine signaling between cells. Our method makes use of the Bio Flip Chip (BFC), a microfabricated silicone chip containing hundreds-to-thousands of microwells, each sized to hold either a single cell or small numbers of cells. We load the chip with cells simply by pipetting them onto the array of wells and washing unloaded cells off the array. The chip is then flipped onto a substrate, whereby the cells fall out of the wells and onto the substrate, maintaining their patterning. After the cells have attached, the chip can be removed (or left on). This approach to cell patterning is unique in that it: 1) doesn't alter the chemistry of the substrate, thus allowing cells to proliferate and migrate; 2) allows patterning onto any substrate, including tissue-culture polystyrene, glass, matrigel, and even feeder cell layers; and 3) is compatible with traditional microcontact printing, allowing the creation of extracellular matrix islands with cells placed inside those islands. In this video, we demonstrate the patterning of mouse embryonic stem cells onto tissue-culture polystyrene using the BFC.     

Microelectrode chip based real time monitoring of vital MCF-7 mamma carcinoma cells by impedance spectroscopy

Sensorchip based impedance spectroscopy can detect inhibitory effects of human neuropeptide Y (hNPY) on living cells in a non-invasive labelling free way in real time without the need of supporting reagents. Since the discovery that neoplasmatic transformations in breast cancer are correlated with a change of the receptor subtype expression of hNPY in the affected tissue, the hNPY receptor-ligand system has come to the fore of cancer research. Today there are different methods detecting hNPY receptor interactions like fluorescent and radioactive labelling or detecting hNPY-pathway activation like cyclic adenosine monophosphate (cAMP) and G protein-coupled receptor (GPCR)-assays. For all these assays it is necessary to either label related proteins with additional substances, which can affect the nature state of the cell, or the need of producing cell lysate which allows only a snapshot of the investigated cells. To overcome these problems we established a new method to detect hNPY-receptor interactions. Therefore, we monitor the complex electric resistance (impedance) of cells attached to a microelectrode over a wide frequency range. Cell alterations are detected as changes in the impedance spectra. After application of the adenylyl cyclase-stimulating reagent forskolin, impedance is decreased at 5kHz frequency within minutes. This effect can be inhibited by preincubating the cells with hNPY for a time range of 20min. The inhibitory effect of hNPY can be washed out and the same cells can be stimulated by forskolin again.     

Single-cell bioelectrical impedance platform for monitoring cellular response to drug treatment

The response of cells to a chemical or biological agent in terms of their impedance changes in real-time is a useful mechanism that can be utilized for a wide variety of biomedical and environmental applications. The use of a single-cell-based analytical platform could be an effective approach to acquiring more sensitive cell impedance measurements, particularly in applications where only diminutive changes in impedance are expected. Here, we report the development of an on-chip cell impedance biosensor with two types of electrodes that host individual cells and cell populations, respectively, to study its efficacy in detecting cellular response. Human glioblastoma (U87MG) cells were patterned on single- and multi-cell electrodes through ligand-mediated natural cell adhesion. We comparatively investigated how these cancer cells on both types of electrodes respond to an ion channel inhibitor, chlorotoxin (CTX), in terms of their shape alternations and impedance changes to exploit the fine detectability of the single-cell-based system. The detecting electrodes hosting single cells exhibited a significant reduction in the real impedance signal, while electrodes hosting confluent monolayer of cells showed little to no impedance change. When single-cell electrodes were treated with CTX of different doses, a dose-dependent impedance change was observed. This enables us to identify the effective dose needed for this particular treatment. Our study demonstrated that this single-cell impedance system may potentially serve as a useful analytical tool for biomedical applications such as environmental toxin detection and drug evaluation.     

Impedance spectroscopy flow cytometry: on-chip label-free cell differentiation.

BACKGROUND: The microfabricated impedance spectroscopy flow cytometer used in this study permits rapid dielectric characterization of a cell population with a simple microfluidic channel. Impedance measurements over a wide frequency range provide information on cell size, membrane capacitance, and cytoplasm conductivity as a function of frequency. The amplitude, opacity, and phase information can be used for discrimination between different cell populations without the use of cell markers. METHODS: Polystyrene beads, red blood cells (RBCs), ghosts, and RBCs fixed in glutaraldehyde were passed through a microfabricated flow cytometer and measured individually by using two simultaneously applied discrete frequencies. The cells were characterized at 1,000 per minute in the frequency range of 350 kHz to 20 MHz. RESULTS: Cell size was easily measured with submicron accuracy. Polystyrene beads and RBCs were differentiated using opacity. RBCs and ghosts were differentiated using phase information, whereas RBCs and fixed RBCs were differentiated using opacity. RBCs fixed using increasing concentrations of glutaraldehyde showed increasing opacity. This increased opacity was linked to decreased cytoplasm conductivity and decreased membrane capacitance, both resulting from protein cross-linking. CONCLUSIONS: This work presents label-free differentiation of cells in an on-chip flow cytometer based on impedance spectroscopy, which will be a powerful tool for cell characterization.     

基于生物阻抗谱的生物细胞活性免标记检测方法

目的 基于生物阻抗谱（bioelectrical impedance spectroscopy,BIS）技术,提出一种免标记的生物细胞活性实时检测方法。该方法依据不同浓度、生理、病理状态下细胞组织的电学特性差异来判断细胞是否具有活性,以协助医师在临床手术中快速精准定位患者烫伤组织并实现有效切除。方法 使用具有活性的斑马鱼胚胎干细胞来模拟人体烫伤组织,采用生物阻抗谱技术来鉴别细胞组织的生理状态。结果 在不同状态下,细胞的阻抗幅值变化有显著的差异,可以从中发现同等浓度下活性细胞的阻抗幅值比死亡细胞平均高出17.25%,活性细胞发生弛豫频率的时间也比死亡细胞早25%。结论 实验数据表明,生物阻抗谱法能有效区分胚胎干细胞的两类生理状态;从聚类区域中可以看出,BIS检测法具有明显的细胞活性及浓度区分能力,理论上能够快速地协助医师完成对患者烫伤组织检测。     

Biosensor for direct cell detection, quantification and analysis

Microarrays are promising tools for cell isolation and detection. However, they have yet to be widely applied in biology. This stems from a lack of demonstration of their sensitivity and compatibility with complex biological samples, and a lack of proof that their use does not induce aberrant cellular effects. Herein, we characterized and optimized a recently developed technology associating antibody microarrays with surface plasmon resonance imaging (SPRi). Using a murine macrophage cell line we demonstrate the binding specificity of our antibody-microarrays and the correlation between SPRi signals and both the number of bound cells, and the level of expression of cell surface markers. Confocal microscopy reveals that cell binding to the chip through antibody-antigen interactions underwent morphological changes reflecting the density of the relevant cell surface marker without affecting cell viability as shown by fluorescent microscopy. The detection threshold of the microarray-SPRi system is lowered 10-fold by applying a polyethylene oxide film to the gold surface of the chip. This increased sensitivity allows the detection of cells representing as little as 0.5% of a mixed population. The potential of this method is illustrated by two applications: characterization of ligand-cell receptor interactions, allowing determination of receptor specificity, and analysis of peripheral blood mononuclear cells, demonstrating the suitability of this tool for the analysis of complex biological samples.     

Integration of single-cell trapping and impedance measurement utilizing microwell electrodes

The ability to research individual cells has been seen as important in many kinds of biological studies. In the present study, cell impedance analysis is integrated into a single-cell trapping structure. For the purpose of precise positioning, a cell manipulation and measurement microchip, which uses an alternating current electrothermal effect (ACET) and a negative dielectrophoresis (nDEP) force to move a particle and cell on measurement electrodes, is developed. An ACET and an nDEP can be easily combined with subsequent analyses based on electric fields. A microwell presented in a previous study is separated into two parts, which are regarded as the measurement electrodes. The original structure is modified for precise positioning. Numerical simulations and analyses are conducted to compute and analyze the effects of the structural parameters. The results of simulations and analyses are used to obtain the optimum structure for the cell. The capture range of the microwell can be designed for cells of various sizes. In order to demonstrate the precision of the positioning, a particle is captured, measured, and released twice. The results show that the impedance error of the particle is about 3%. Finally, the developed structure is applied to trap and measure the impedance of a HeLa cell.     

Comparison of impedance-based sensors for cell adhesion monitoring and in vitro methods for detecting cytotoxicity induced by chemicals

The recent European Commission REACH (Registration, Evaluation and Authorisation of Chemicals) policy outlines a plan for toxicological testing by using alternative non-animal in vitro methods. In this context, there is a need to develop and standardise high-throughput screening (HTS) methods for studying the cytotoxicity induced by chemicals. Electrochemical impedance spectroscopy (EIS) can be considered as a complementary technique to alternative in vitro testing for studying cell adhesion to the substrate, and can give real-time and kinetic information on cell responses to a toxicant. This paper describes the development of a home-made chip based on impedance spectroscopy, and its application in studying the kinetics of BALB/3T3 cell adhesion and the cellular responses to a toxic product as a function of time. Concentrations of sodium arsenite, ranging from 10 microM up to 1000 microM, were tested in the system, and the results were compared with those obtained with standard protocols used to study basal cytotoxicity induced by chemicals in the BALB/3T3 cell line. The results show that the sensitivity of the developed chip was better than that with the MTT test, with the additional advantages of online monitoring.     

On-chip non-invasive and label-free cell discrimination by impedance spectroscopy

OBJECTIVES: Many flow-cytometric cell characterization methods require costly markers and colour reagents. We present here a novel device for cell discrimination based on impedance measurement of electrical cell properties in a microfluidic chip, without the need of extensive sample preparation steps and the requirement of labelling dyes. MATERIALS AND METHODS, RESULTS: We demonstrate that in-flow single cell measurements in our microchip allow for discrimination of various cell line types, such as undifferentiated mouse fibroblasts 3T3-L1 and adipocytes on the one hand, or human monocytes and in vitro differentiated dendritic cells and macrophages on the other hand. In addition, viability and apoptosis analyses were carried out successfully for Jurkat cell models. Studies on several species, including bacteria or fungi, demonstrate not only the capability to enumerate these cells, but also show that even other microbiological life cycle phases can be visualized. CONCLUSIONS: These results underline the potential of impedance spectroscopy flow cytometry as a valuable complement to other known cytometers and cell detection systems.     

On-line monitoring of lipid storage in yeasts using impedance spectroscopy

Bioremediation technologies and many environmentally sound biosyntheses rely on the catalytic potential of whole cells. For analyzing and controlling such processes robust real-time indicators for the concentration of intact cells such as impedance are required. The conventional method measures the capacitances of cell suspensions at one or two frequencies and correlates them with biomass concentrations. However, cell inclusions such as lipid droplets or overproduced enzymes may block intracellular ion paths, thereby possibly modifying the dielectric properties of the cells. To test the hypothesis that the total impedance spectrum into the analysis may provide useful information about cell inclusions, the impedance spectrum of a technical culture of the oleaginous yeast Arxula adeninivorans was measured and evaluated every 15 s. This yeast is a good test object since it stores the excess of assimilated carbon in experimentally controllable lipid droplets. Upon correction for possible impedance signal interferences, we derived different empirical methods suitable to indicate incipient lipid formation. The methods were designed to act on-line and are thus principally suited for real-time monitoring of cell inclusions. In search for optimised bioprocess monitoring we tested a heuristic spectrum analysis using integrative statistics (RDA). With this approach we were able to accurately detect the formation of cell inclusions, which is potentially valuable for future bioprocess control strategies.     

Microcavity array (MCA)-based biosensor chip for functional drug screening of 3D tissue models

Multicellular tumour spheroids that mimic a native cellular environment are widely used as model systems for drug testing. To study drug effects on three-dimensional cultures in real-time we designed and fabricated a novel type of sensor chip for fast, non-destructive impedance spectroscopy and extracellular recording. Precultured spheroids are trapped between four gold electrodes. Fifteen individual 100microm deep square microcavities with sizes from 200 to 400microm allow an optimised positioning during the measurement. Although apoptosis was induced in human melanoma spheroids by Camptothecin (CTT), treated cultures did not show disintegration but displayed increased impedance magnitudes compared to controls after 8h resulting from an altered morphology of the outer cells. Contractions in cardiomyocyte spheroids were monitored when the innovative chip was used for recording of extracellular potentials. The silicon-based electrode array is used as an acute test system for the monitoring of any kind of 3D cell cultures. Since no adherence of cells or labelling is necessary the multifunctional sensor chip provides a basis for improved drug development by high content screenings with reduced costs and assay times. Additional improvements for parallel testing of different substances on one chip are presented.     

Cell-based chip for the detection of anticancer effect on HeLa cells using cyclic voltammetry

HeLa cells directly immobilized on gold-patterned silicon substrate were used to assess the biological toxicity of anticancer drugs (hydroxyurea and cyclophosphamide). Immobilization of HeLa cells was confirmed by optical microscopy, and cell growth, viability and drug-related toxicity were examined by cyclic voltammetry and potentiometric stripping analysis. The voltammetric behaviors of HeLa cells displayed a quasi-reversible pattern with the peak current exhibiting a linear relationship with cell number. The attached living cells were exposed to different concentrations of hydroxyurea and cyclophosphamide as anticancer drugs, which induced the change of cyclic voltammetry current peak. As the exposed concentration of anticancer drugs was increased, the change of current peak was increased, which indicates the decrease of cell viability. Trypan Blue dyeing was performed to confirm the results of the effect of anticancer drugs on the cell viability which was obtained from cyclic voltammetry assay. The proposed direct cell immobilization method technique can be applied to the fabrication of cell chip for diagnosis, drug detection, and on-site monitoring.     

Real-time detection of β1 integrin expression on MG-63 cells using electrochemical impedance spectroscopy

Beta 1 integrin is a membrane protein responsible for attachment and migration of osteosarcoma cells. In this study, expression of β1 integrin on MG-63 cells, a human osteogenic sarcoma cell line, was monitored using electrochemical impedance spectroscopy (EIS). ITO-based biochips were developed using a semiconductor technique. Differences in electric resistance (ΔR) were measured continuously when cells binding with anti-β1 integrin antibody coagulated with nano-scale gold particles. The results of the EIS system were compared with traditional immunofluorescence staining. We found that sample chambers with higher cell densities had larger ΔR values. When the cell densities increased from 5 × 10(4) cells/ml to 5 × 10(5) cells/ml, the ΔR value dose-dependently increased from 14 Ω to 37 Ω. In addition, a highly linear relationship (correlation coefficient, 0.921) was found between the ΔR values and the corresponding fluorescence intensities (p<0.05). These results suggest that electrochemical impedance spectroscopy can be a useful tool for evaluating β1 integrin expression on cell membranes.