Theoretical modelling of interdigitated electrode sensor for mammalian cell characterization

Interdigitated Electrodes (IDEs) have been widely used in biological cellular characterization such as the Electrical Cell-Substrate Impedance Sensing (ECIS). Optimization of IDEs are crucial to obtain high accuracy of measurement that associates with the biological cell activities. However, not much research studies the generation of electric field by the IDEs geometry especially in cellular application. In this work, theoretical modelling of IDEs was done by modelling the IDEs equivalent circuit consisting of 3 major components; double layer capacitance, CDL, solution capacitance, CSOL and solution resistance, RSOL. Simulation using MATLAB and COMSOL Multiphysics was done to study the effect of geometrical parameters (width of electrodes (W), spacing between electrodes (S) and total number of electrodes (N)) on the cut-off frequency (FLOW), solution resistance (RSOL) and the average electric field magnitude based on the equivalent circuit model. The simulation results show three main findings; lowest FLOW to be at the ratio of a=0.54 and N16, lowest RSOL at smaller a and higher N, and saturated electric field at N18. The results suggested that the optimal configuration of IDEs with a fixed length of electrode of 7000μm is to have the ratio of (S/W) as 0.54 and N as 18

Cytotoxicity studies of lung cancer cells using impedance biosensor

Electrical cell-substrate impedance sensing (ECIS) is a valuable tool for real time monitoring of cell behavior such as attachment, mobility, and growth. To employ ECIS, the cells need to attach, spread and proliferate on the sensor in the presence of adhesion-promoting protein that mimics the extracellular matrix (ECM) of the cells. For cell attachment, collagen I, Bovine had been used as the coating substrate. In this study, four designs with varying electrode distances had been measured to detect the changes in impedance values of Lung Carcinoma cell lines (A549). The impedance change due to the cell growth and attachment was modeled as an equivalent circuit consisting of resistors and capacitors of both the cell culture media and the cells. The impedance measurements were measured every 8 hours for 120 hours at frequencies of 100Hz to 10MHz using Agilent Precision Impedance Analyzer 4294A. The experimental results have shown that the closest distance of the electrode gave the most optimum impedance value for A549 cancer cell’s measurement. The cancer cells were also treated with a chemotherapeutic drug, Taxol and its impedance response was monitored over 5 days. Experimental results show that there is significant reduction in impedance when the cancer cells were exposed to Taxol, indicating that the cells are no longer adherent to the sensor’s surface or are dead

Modeling and development of screen-printed impedance biosensor for cytotoxicity studies of lung carcinoma cells

Electrical cell-substrate impedance sensing (ECIS) is a powerful technique to monitor real-time cell behavior. In this study, an ECIS biosensor formed using two interdigitated electrode structures (IDEs) was used to monitor cell behavior and its response to toxicants. Three different sensors with varied electrode spacing were first modeled using COMSOL Multiphysics and then fabricated and tested. The silver/silver chloride IDEs were fabricated using a screenprinting technique and incorporated with polydimethylsiloxane (PDMS) cell culture wells. To study the effectiveness of the biosensor, A549 lung carcinoma cells were seeded in the culture wells together with collagen as an extracellular matrix (ECM) to promote cell attachment on electrodes. A549 cells were cultured in the chambers and impedance measurements were taken at 12-h intervals for 120 h. Cell index (CI) for both designs were calculated from the impedance measurement and plotted in comparison with the growth profile of the cells in T-flasks. To verify that the ECIS biosensor can also be used to study cell response to toxicants, the A549 cells were also treated with anti-cancer drug, paclitaxel, and its responses were monitored over 5 days. Both simulation and experimental results show better sensitivity for smaller spacing between electrodes

Screen printed impedance biosensor for cytotoxicity studies of lung carcinoma cells

Electrical Cell-Substrate Impedance Sensing is a powerful tool for monitoring real time cells properties such as adhesion, mobility and cytotoxicity. In this study, a silver/silver chloride screen-printed impedance biosensor was developed to characterize A549 lung carcinoma cells growth in the presence of collagen I, Bovine. Collagen acts as an extracellular matrix (ECM) for A549 and promotes cell attachment. The sensor was incorporated with a culture well which was fabricated from polydimethylsiloxane (PDMS). A549 cells were cultured in the chambers and impedance measurements were taken at 12 hours intervals for 120 hours. Cell Index (CI) were calculated from the impedance data and plotted in comparison with growth profile of the cells in T-flasks for validation of the sen- sor’s functionality. A549 cells were also treated with anti- tumor drug; Paclitaxel and its response were monitored over 5 days. Experimental results show significant change in CI dur- ing growth and death after exposure to Taxol, indicating that tumor growth was inhibited in the presence of Taxol

Toxicity studies of agarwood essential oil in vero cells using electrical impedance sensor

Natural products have traditionally been used for medicinal purposes in Asian communities. Toxicity studies typically use animal testing to predict the harmfulness of a particular substance to human health. For this study, in lieu of animal testing, we utilized cell-based biosensors to evaluate the toxicity of natural products. The cell-based biosensors were fabricated on a printed circuit board with copper electrodes and equipped with PDMS cell culture chambers. Two different electrodes (interdigitated and circular) were designed. Vero cells were used to represent normal healthy cells. The cells were first cultured on the biosensors and then inoculated with natural products, Taxol (chemo drug – positive control), and DMSO (negative control). Impedances of these biosensors were afterwards recorded at six-hour intervals for 80 hours to determine the growth of the cells. It was found that as compared to Taxol, natural products have substantially low toxicant values

Integrated multichannel electrochemical–quartz crystal microbalance sensors for liquid sensing

This paper highlights the design, simulation and fabrication of an array of twelve integrated electrochemical – quartz crystal microbalance (IEQCM) sensors on a single substrate for liquid sensing. Integration of both measurement techniques is made possible by combining the three electrode electrochemical device with the top and bottom electrodes for the microbalance. Important design parameters such as the working electrode radius and gap spacing, were studied using both theoretical calculations and COMSOL Multiphysics R finite element simulations. The sensor’s working electrode radius affects the magnitude of the frequency response while the gap affects the capacitance and current density which are important for electrochemical measurements. It was found that the best values for the working electrode radius was 2 mm and gap spacing was 0.5 mm. The sensors were fabricated using microfabrication techniques for the gold electrode and screen printing techniques for the reference electrode. Water contact angle, atomic force microscopy, and scanning electron microscope were utilized to study the surface roughness of the IEQCM sensor. IEQCM has a low contact angle of 53.0 ± 1 ◦ and low surface roughness of 1.92nm. For liquid sensing, an array of circular chambers were fabricated using polydimethylsiloxane (PDMS) and placed on top of the quartz substrate for liquid testing. Electrochemical measurements and cyclic voltammetry were performed using the sensor in ferri-ferrocyanide and phosphate buffered saline solution to study the function of scan rates on the peak current with respect to the potential difference. For mass sensing measurements, liquid water droplets of 1uL – 10 uL were placed onto the sensing surface and the change in resonance frequencies of the sensors were measured. These resonance frequency changes can be converted in mass change/area in accordance to the advanced Sauerbrey equation. The multichannel IEQCM sensor shows good potential as a parallel sensor for both biosensing and environmental applications

Simulation of ring interdigitated electrode for dielectrophoretic trapping

Electric field intensity is important in trapping biological cells during dielectrophoresis (DEP). In this paper, two designs of ring interdigitated electrode (RIDE) with varied spacing between electrodes; 300 µm to 500 µm, were modelled and analyzed. Analysis was done using finite element analysis software, COMSOL Multiphysics to study the intensities of electric fields generated on the electrodes. Simulation results show that higher electric fields are generated by the asymmetrical RIDE compared to the symmetrical RIDE design. The average value of positive electric fields peaks for symmetrical RIDE is 16.1kV/m and 19.9 kV/m for asymmetrical RIDE. Simulations also revealed that higher electric field were generated on smaller spacing compared to larger one. This suggested that better cellular attraction can be predicted on smallest distance of asymmetrical RIDE. Trapped cells can later be used to study the intercellular or intracellular interactions of the specific cells, such as through impedance sensing to form an integrated DEP-impedance biosensor

Optimization of printing techniques for electrochemical biosensors

Electrochemical biosensors show great promise for point-of-care applications due to their low cost, portability and compatibility with microfluidics. The miniature size of these sensors provides advantages in terms of sensitivity, specificity and allows them to be mass produced in arrays. The most reliable fabrication technique for these sensors is lithography followed by metal deposition using sputtering or chemical vapor deposition techniques. This technique which is usually done in the cleanroom requires expensive masking followed by deposition. Recently, cheaper printing techniques such as screen-printing and ink-jet printing have become popular due to its low cost, ease of fabrication and mask-less method. In this paper, two different printing techniques namely inkjet and screen printing are demonstrated for an electrochemical biosensor. For ink-jet printing technique, optimization of key printing parameters, such as pulse voltages, drop spacing and waveform setting, in-house temperature and cure annealing for obtaining the high quality droplets, are discussed. These factors are compared with screen-printing parameters such as mesh size, emulsion thickness, minimum spacing of lines and curing times. The reliability and reproducibility of the sensors are evaluated using scotch tape test, resistivity and profile-meter measurements. It was found that inkjet printing is superior because it is mask-less, has minimum resolution of 100 μm compared to 200 μm for screen printing and higher reproducibility rate of 90% compared to 78% for screen printing

Microfluidic concentration gradient for toxicity studies of lung carcinoma cells

Cancer is a serious global health problem, which resulted in 8.2 million deaths in 2012 alone. Amongst different types of cancer, lung cancer is the most lethal and contributes 19.4% of cancer deaths. Better disease-free cancer survival rates have been reported when surgery is followed by systemic chemotherapy. Efficient treatment can be achieved through personalized chemotherapy dosage whereby optimum treatment is given to kill the cancer the side effects are minimized. Here, we present a microfluidic concentration gradient device for toxicity studies on lung cancer cell lines (A549). Automated drug dilution is achieved by simply tuning the flow rate and geometries of the microfluidics network. Sets of tree-like-concentration-generators were designed to achieve constant flow rate at each outlet by optimizing the channel lengths. Serpentine structures were placed in the middle in the middle and at each outlet channel to the design to improve mixing along the channel. The lengths of middle and outlet channels are varied from 1.5 mm to 12 mm to obtain sufficient mixing of two fluid flows. Theoretically, correlations between hydraulic flow and electrical circuit equations analogy were applied to ease the microfluidic design process. Later, 3D (dimensional) simulations using computational fluid dynamic (CFD)-based simulator, i.e. Ansys FLUENT were performed by implementing species transport method prior to fabrication. The simulation process helps to demonstrate the effect of varying channel length on the velocity magnitude and the concentration of the microfluidic structure. In addition, the simulation results allows us predict the fluid flow velocity that showed constant velocity magnitude at each outlet. Wider range of dilution can be achieved, when a higher number of outlets are added in a microfluidic design. Polydimethylsiloxane (PDMS) microchannels were fabricated on glass slide widths of 200 μm and depths of 35 μm using soft-lithography technique [1]. The 3-outlet serpentine structure produced the best match between simulation and measurement results. The concentration profiles produce inside the channel is determined by the splitting ratio of the fluids at each branched and also depends on the number of the inlet and outlet in the tree-like network. The gradient generator will be attached to an array of cell culture chambers with sensors that were previously developed for toxicity studies of lung cancer (A549) cell lines is shown in the Fig. 2. Cells cultured in the sensor will begin to attach and spread on the surface of the electrodes, restricting current flows from the electrodes to the surrounding media. In a confluent (all surface covered with cells) cell layer, current must travel through the intercellular space of the cell-cell and also the tight gap of the cell-electrode pairs to reach surrounding media. The more adhered the cells are with each other and with the electrode, the lesser the amount of current that could travel out, thus increasing the overall impedance of the system. This leads to a good way of studying cell-cell and cell-electrode adhesion characteristics by using impedance measurement [2] ; [3]. When sensors are treated with Taxol, the cell index (CI) values of the cancer cells exhibit inconsistent trend with several peaks during the measurement (over 96 hours) as shown in Fig.1. This is due to the nature of cells that are mixed combinations of drug-sensitive cells and drug resistance cells. This work provides a promising solution for automated drug dilution in parallel toxicity studies. The use of microfluidics allows highly parallel, maximum testing with minimal reagents to obtain the optimum dosage

Content cytotoxicity studies of colorectal carcinoma cells using printed impedance sensors

Monitoring the effectiveness of drugs on cancer cells is crucial for chemotherapeutics studies. Invitro cell-based biosensors can be used as an alternative for characteristic studies of cells’ response to drugs. Cell-based sensors provide real-time measurements and require smaller sample volumes compared to conventional T-flask measurement methods. This paper presents a biosensor that detects in real-time, impedance variations of human colon cancer, HCT-116 cells when treated with anti-cancer agent, 5-Fluorouracil (5-FU). Two different extracellular matrix (ECM); polyaniline and gelatin were tested and evaluated in terms of attachment quality. Polyaniline was found to provide the best attachment for HCT-116 cells and was used for cytotoxicity studies. Cytokinetic behavior indicated that 5-FU inhibited HCT-116 cells at IC50 of 6.8 µg/mL. Trypan blue exclusion method for testing cell viability was used to validate the impedance measurements, where the cancer cell concentrations were reduced to ~35% when treated with 2.5 µg/mL, and 50% when treated with 6.8 µg/mL. The results generated by the microfabricated impedance biosensor are comparable to the Trypan blue method since both gave similar cell growth trend. It can be concluded that the impedance biosensor has potential to be used as an alternative method in drug testing application