Electrical Biosensors Research Articles

Detection of bacterial metabolism in lag-phase using impedance spectroscopy of agar-integrated 3D microelectrodes

Traditional methods for detection of metabolically-active bacterial cells, while effective, require several days to complete. Development of sensitive electrical biosensors is highly desirable for rapid detection and counting of pathogens in food, water, or clinical samples. Herein, we develop a highly-sensitive non-Faradaic impedance sensor which detects metabolic activity of E. coli cells in a mere 1 μl of sample volume and without any sample filtration/purification. The three dimensional (3D) interdigitated electrodes (IDEs) along with self-assembled gold-nickel (Au-Ni) nanostructures significantly amplify the sensitivity by increasing the sensing area almost three-fold. The developed microsystem is integrated with an agar-based growth medium and monitors the metabolism of bacterial cells, enabling bacterial detection in approximately one hour after inoculation, i.e. in the lag-phase. Incorporation of a secondary agar layer as a biocompatible passivation layer protects the IDEs from potential Faradaic reactions and enhances sensitivity to modulation of the non-Faradaic impedance due to cellular metabolism. The resultant label-free sensor is capable of selective identification of metabolizing cells (vs. dead cells) across a wide linear range (10–1000 cells/μl). These results help pave the way for rapid antibacterial susceptibility testing at the point-of-need, which is currently a major challenge in healthcare.

Field-Effect Transistor-Integration with TiO2 Nanoparticles for Sensing of Cardiac Troponin I Biomarker.

The development of electrical biosensor towards device miniaturization in order to achieve better sensitivity with enhanced electrical signal has certain limitations especially complexity in fabrication process and costs. In this paper, an alternative technique with minor modification in the device structure is presented for signal amplification by implementing ambipolar conduction in the biosensor itself. We demonstrated the field-effect transistor (FET)-based biosensor coupled back-gate for attaining a higher sensitivity with the detection of lower target abundance. To utilize the coupled back-gate as a pre-amplifier, silicon-on-insulator wafer with thicknesses of top-silicon and buried oxide (BOX) layers of 70 nm and 145 nm, respectively were desired. Titanium dioxide (TiO2) nanomaterial was deposited using sol-gel method on the channel which acts as a transducer. Surface functionalization on TiO2 thin film allowed an effective immobilization of anti-cardiac troponin I antibody to interact cardiac troponin I (cTnI). Binding events at each step was validated by X-ray photoelectron spectroscopy (XPS) analysis. Further, electrical characterization (Id-Vd) confirms the potentiality of FET-based biosensor to detect cTnI (represents acute myocardial infarction disease) with the concentration ranges from 10 μg/ml down to 1 fg/ml. The sensitivity of 459.2 nA (g/ml)-1 and lower detection limit of 1 fg/ml were achieved at Vbg = -5 V and Vd = 5 V. The designed device demonstrates its ability to detect lower level of cTnI with pre-amplified electrical signal by back-gate biasing.

High-performance nanogap electrode-based impedimetric sensor for direct DNA assays

The rapid and sensitive detection of pathogen DNA (Deoxyribonucleic acid) would be essential for diagnosis and appropriate antibiotic treatment time. Herein, we report a novel direct DNA detectable impedimetric sensor. Direct assay of the amplified target DNA (mecA gene from methicillin-resistant Staphylococcus aureus (MRSA)) was performed using the PCR (polymerase chain reaction) product without any purification. Even though there are lots of PCR reagents and excess salts in sample PCR product, the nanogap electrode-based impedimetric sensor was able to detect DNA amplification fast in 5th PCR cycle which had 260 fM mecA gene in sample originally. The 70 nm gap electrode sensor yielded over 20% signal increase at the 5th PCR cycle and the impedance change grew up to about 60% at 25th in case of sample with 260 fM mecA gene template originally. The increased concentration of target DNA template led to the rise in impedance change such as 60% up at 5th and 120% up at 25th cycle with 260 pM, respectively. It is very outstanding result as compared with the traditional PCR agarose gel. Besides, it is 7-fold superior sensitivity to the microgap electrode. Furthermore, genomic DNA sample extracted from MRSA was detected rapidly. The nanogap electrode-based impedimetric sensor could be a good candidate for a rapid, sensitive, and low-cost electrical biosensor for DNA characterization in diagnostics and disease monitoring.

Graphene Nanoporous FET Biosensor: Influence of Pore Dimension on Sensing Performance in Complex Analyte

Recently, there have been efforts to realize ultra-sensitive electrical biosensors on 3-D graphene nanostructures like nanowalls or nanopores which can enhance the surface area to volume ratio. But, the signal generated from the nanowall/nanopore structures for low concentration of biomolecules is often insufficient for current measurements and might not be reliable, especially for complex physiological analyte. This is the major drawback with respect to practical deployment of such sensors. In this paper, we have explored the impact of pore morphology on the sensing performance of label free, non-faradaic graphene nanoporous field effect biosensor with a view to achieve sub-femtomolar detection limit with an appreciable signal-to-noise ratio (SNR) in serum. Nine different pore geometries have been fabricated and the sensors have been experimented with Hep-B and Hep-C as specific and non-specific antigen, respectively. It has been observed that the graphene sensor with pore diameter of 35 nm and length of 160 nm has the maximum SNR in serum, in spite of not having the highest surface area to volume ratio and hence in nanoporous graphene structure, design of pore morphology should not be inclined towards increasing the surface area alone. The detection limit obtained with the optimized geometry is 0.1-fM Hep-B in a complex mixture of serum containing 1nM Hep-C as non-specific antigen.

Multifunctionalized Reduced Graphene Oxide Biosensors for Simultaneous Monitoring of Structural Changes in Amyloid-β 40.

Determination of the conformation (monomer, oligomer, or fibril) of amyloid peptide aggregates in the human brain is essential for the diagnosis and treatment of Alzheimer’s disease (AD). Accordingly, systematic investigation of amyloid conformation using analytical tools is essential for precisely quantifying the relative amounts of the three conformations of amyloid peptide. Here, we developed a reduced graphene oxide (rGO) based multiplexing biosensor that could be used to monitor the relative amounts of the three conformations of various amyloid-β 40 (Aβ40) fluids. The electrical rGO biosensor was composed of a multichannel sensor array capable of individual detection of monomers, oligomers, and fibrils in a single amyloid fluid sample. From the performance test of each sensor, we showed that this method had good analytical sensitivity (1 pg/mL) and a fairly wide dynamic range (1 pg/mL to 10 ng/mL) for each conformation of Aβ40. To verify whether the rGO biosensor could be used to evaluate the relative amounts of the three conformations, various amyloid solutions (monomeric Aβ40, aggregated Aβ40, and disaggregated Aβ40 solutions) were employed. Notably, different trends in the relative amounts of the three conformations were observed in each amyloid solution, indicating that this information could serve as an important parameter in the clinical setting. Accordingly, our analytical tool could precisely detect the relative amounts of the three conformations of Aβ40 and may have potential applications as a diagnostic system for AD.

Low Cost PCB Fuel Cells Based for Small Electronic Applications

An important aspect to enhance the introduction of the fuel cells systems in the market, is decreasing the cost of manufacturing, obtaining a flexible way to extend their use in different applications. The possibility to use the well-defined PCB manufacturing technic to develop fuel cells stacks can help to obtain a low-cost and tuneable method for their production [1]. Using this manufacturing method is easy to control the size and adjust the power requirements for different electronic devices as electrochromic devices, smartphone chargers, electric vehicles, etc. As well, due to their flat configuration is very easy to stack many of them in small spaces. In this work we design and develop a mini and micro stack DMFC open cathode PCB cell using low cost materials on the bipolar plates and electrodes. In general, for the bipolar plates different metal and ink coatings were used to improve the durability and the electrical contact. But them requires few steps to obtain the final stage, in our case we moved to a graphitic flexible material with very high corrosion resistance and conductivity giving the possibility to avoid the coating steps. In the case of the electrodes, a non-precious metal and very high poison resistance catalyst developed in our lab was used as cathode [2]. In the anode part, an inkjet printed electrodes where used. This method gives the possibility to decrease the loading of catalyst obtaining a really good performance in the Fuel Cell. For the micro-DMFC we developed two setups, one similar to the mini stack cell and the other one, in a complete passive configuration. The design of the fully passive cells gives the possibility to inject the methanol solution in the anode reservoirs. This micro stack cell was developed to power up an autonomous electrical biosensor synergistically assembled inside a passive DMFC for screening cancer biomarkers included in the European 2020 project called Symbiotic. [1] O. A. Obeisun et al., “Development of open-cathode polymer electrolyte fuel cells using printed circuit board flow-field plates: Flow geometry characterisation,” Int. J. Hydrogen Energy, vol. 39, no. 32, pp. 18326–18336, 2014. [2] D. Malko, T. Lopes, E. Symianakis, and A. R. Kucernak, “The intriguing poison tolerance of non-precious metal oxygen reduction reaction (ORR) catalysts,” J. Mater. Chem. A, vol. 4, no. 1, pp. 142–152, 2016. Figure 1

Neuropeptide oxytocin enhances μ opioid receptor signaling as a positive allosteric modulator

Oxytocin (OT) is a 9-amine neuropeptide that plays an essential role in mammalian labor, lactation, maternal bonding, and social affiliation. OT has been reported to exert an analgesic effect in both humans and animals, and the results of certain animal experiments have shown that the analgesic effect of OT is partially blocked by opioid receptor antagonists. To investigate the relationship between OT and μ opioid receptor (MOR), we evaluated how OT affects MOR in vitro by performing an electrical impedance-based receptor biosensor assay (CellKey™ assay), an intracellular cAMP assay, and a competitive receptor-binding analysis by using cells stably expressing human MOR and OT receptor. In both the CellKey™ assay and the intracellular cAMP assay, OT alone exerted no direct agonistic effect on human MOR, but treatment with 10−6 M OT markedly enhanced the MOR signaling induced by 10−6 M endomorphin-1, β-endorphin, morphine, fentanyl, and DAMGO. Moreover, in the competitive receptor-binding assay, 10−6 M OT did not alter the affinity of endomorphin-1 or morphine for MOR. These results suggest that OT could function as a positive allosteric modulator that regulates the efficacy of MOR signaling, and thus OT might represent a previously unrecognized candidate analgesic agent.

Advances in Nanowire Transistor‐Based Biosensors

AbstractSemiconducting nanowires (NWs) present new opportunities for developing electrical biosensors due to their inherent properties, including large surface‐to‐volume ratio, rapid signal response, configurability in electronic devices, and nanoscale footprint comparable to biomolecular and subcellular structures. Field‐effect transistors (FETs) based on silicon nanowires (SiNWs) have proven to be a powerful and versatile tool in the biomedical fields. Recent advances in SiNW‐FET biosensors, including fundamental aspects and working principles, preparation of nanowires, and surface functionalization, as well as applications in biological analysis and cellular investigation, are discussed here. It is envisioned that nano‐FET devices will have several applications in fields such as clinical diagnosis, point‐of‐care testing, and screening for early disease.

Dynamic monitoring of transmembrane potential changes: a study of ion channels using an electrical double layer-gated FET biosensor.

In this research, we have designed, fabricated and characterized an electrical double layer (EDL)-gated AlGaN/GaN high electron mobility transistor (HEMT) biosensor array to study the transmembrane potential changes of cells. The sensor array platform is designed to detect and count circulating tumor cells (CTCs) of colorectal cancer (CRC) and investigate cellular bioelectric signals. Using the EDL FET biosensor platform, cellular responses can be studied in physiological salt concentrations, thereby eliminating complex automation. Upon investigation, we discovered that our sensor response follows the transmembrane potential changes of captured cells. Our whole cell sensor platform can be used to monitor the dynamic changes in the membrane potential of cells. The effects of continuously changing electrolyte ion concentrations and ion channel blocking using cadmium are investigated. This methodology has the potential to be used as an electrophysiological probe for studying ion channel gating and the interaction of biomolecules in cells. The sensor can also be a point-of-care diagnostic tool for rapid screening of diseases.

Current Developments and Potential Applications of Biosensor Technology

The development of various new kinds of sensors for the accurate detection of biomarkers in biological fluids and environmental samples are of greatest importance for the early diagnosis of diseases and to avoid the contamination of environment through pollutants, toxic and biohazardous materials. Sensitivity limits of biosensor have increased due to developments of new biological methods like tagging of fluorescence molecule with nanomaterials. Moreover, usage of peptide arrays, aptamers, antibodies, nucleotides and molecule fixed polymers, facilitate to improve advanced biosensors over conventional approaches. Several biosensors ranging from nanomaterials, polymers to microbes have broader potential applications. Generally, biosensor has been organized into several categories containing diverse sensing arrangements such as mechanical, optical and electrical transducers and modern biosensors use micro- and nanofabrication tools, as either label-free or labeled. This review provides an overview of recent developments and applications of biosensors in the fields of biomedical sciences and environmental monitoring, along with the better detection limit and improved sensitivity of the biosensors.

Detection of prostate specific antigen (PSA) in human saliva using an ultra-sensitive nanocomposite of graphene nanoplatelets with diblock-co-polymers and Au electrodes.

Prostate-specific antigen (PSA) is a commonly used biomarker for the detection of prostate cancer (PCa) and there are numerous data available for its invasive detection in the serum and whole blood. In this work, an electrochemical sensing method was devised to detect traces of PSA in human saliva using a hybrid nanocomposite of graphene nanoplatelets with diblock co-polymers and Au electrodes (GRP-PS67-b-PAA27-Au). The pure graphitic composition on filter paper provides significantly high electrical and thermal conductivity while PS67-b-PAA27 makes an amphiphilic bridge between GRP units. The sensor utilizes the binding of an anti-PSA antibody with an antigen-PSA to act as a resistor in a circuit providing an impedance change that in turn allows for the detection and quantification of PSA in saliva samples. A miniaturized electrical impedance analyzer was interfaced with a sensor chip and the data were recorded in real-time using a Bluetooth-enabled module. This fully integrated and optimized sensing device exhibited a wide PSA range of detection from 0.1 pg mL-1 to 100 ng mL-1 (R2 = 0.963) with a lower limit of detection of 40 fg mL-1. The performance of the biosensor chip was validated with an enzyme-linked immunosorbent assay technique with a regression coefficient as high as 0.940. The advantages of the newly developed saliva-PSA electrical biosensor over previously reported serum-PSA electrochemical biosensors include a faster response time (3-5 min) to achieve a stable electrical signal for PSA detection, high selectivity, improved sensitivity, no additional requirement of a redox electrolyte for electron exchange and excellent shelf life. The presented sensor is aimed for clinical commercialization to detect PSA in human saliva.

High Performance Biosensor Based on Electronic Transduction of Refractive Index Using 3D Nanostructured Biogenic Silica

Various nanostructured materials have been deployed in developing biosensors. Nevertheless, achieving sub-attomolar detection limit by label free method is still challenging owing to the different noises during the sensing process. In this paper, we introduce an impedance immunosensor on freely suspended 3D nanostructured biogenic silica block, commonly called Tabashir. The working principle of this sensor is based on the electrical transduction of variations in refractive index after antigen capture. Due to the variation in refractive index, the Tabashir sensor changes the solution lining on the surface and within the pores, which has been converted to a significant variation in bulk conductivity. This screens the impedance measurement from major noise interference like the surface charge fluctuations and antibody-antigen association dissociation transients, resulting in a remarkable steady state impedance sensitivity of around 120% along with distinct shape and cut-off frequencies in the noise power spectral density (PSD) for a trace concentration of 0.1 aM Hep-B virus even in a complex mixture containing high concentration of non-specific antigen. This detection limit is an improvement by almost two orders of magnitude compared to the most sensitive label free electrical biosensor.

ScFv-modified graphene-coated IDE-arrays for ‘label-free’ screening of cardiovascular disease biomarkers in physiological saline

Fatty-acid binding proteins (FABP) and myeloperoxidases (MPO) are associated with many chronic conditions in humans and considered to be important biomarkers for diagnosis of cardiac diseases. Here we assemble a new electrical biosensor platform based on graphene-coated interdigitated electrode arrays (IDE-arrays) towards ultrafast, label-free screening of heart type-FABP and MPO. Arrays of nanoscale (nanoIDE) and microscale (microIDE) electrode-arrays were fabricated on wafer-scale by combining nanoimprint and photolithography processes. Chemical vapor deposition grown multilayer graphene was transferred onto nano/microIDE-arrays and used as a high surface-to-volume ratio electrical transducer. Novel biofunctional layers of specially engineered anti-h-FABP and anti-MPO single-chain fragment variables (scFv) were immobilized onto graphene-coated IDE-array sensor platform for electrical detection of h-FABP and MPO in physiological saline. scFv fragments show increased sensitivity in comparison to the state-of-the-art competitive ELISA for their higher affinity towards target analytes. Deploying FABP and MPO specific scFvs as receptor molecules onto our high-sensitivity graphene-coated IDE-arrays with identical sensor characteristics and assays covering clinically relevant concentrations in physiological saline, we demonstrate realization of a simple and versatile biosensor platform capable of high performance cardiac-bioassays for point-of-care applications.

Fabrication and Characterization of a Stabilized Thin Film Ag/AgCl Reference Electrode Modified with Self-Assembled Monolayer of Alkane Thiol Chains for Rapid Biosensing Applications.

The fabrication of miniaturized electrical biosensing devices can enable the rapid on-chip detection of biomarkers such as miRNA molecules, which is highly important in early-stage cancer detection. The challenge in realizing such devices remains in the miniaturization of the reference electrodes, which is an integral part of electrical detection. Here, we report on a novel thin film Ag/AgCl reference electrode (RE) that has been fabricated on top of a Au-sputtered glass surface, which was coated with a self-assembled monolayer (SAM) of 6-mercepto-1-hexanol (MCH). The electrode showed very little measurement deviation (−1.5 mv) from a commercial Ag/AgCl reference electrode and exhibited a potential drift of only ± 0.2 mV/h. In addition, the integration of this SAM-modified microfabricated thin film RE enabled the rapid detection (<30 min) of miRNA (let-7a). The electrode can be integrated seamlessly into a microfluidic device, allowing the highly stable and fast measurement of surface potential and is expected to be very useful for the development of miniature electrical biosensors.

Self-Assembled DNA Generated Electric Current Biosensor for HER2 Analysis.

We have developed a new DNA self-assembly amplification technology that generates electric current for electrochemical biosensing. The new technology was used for detection of human epidermal growth factor receptor 2 (HER2). In our technology, an aptamer was utilized both as a ligand for recognition and as a signal generating reporter. The aptasensor is based on a sandwich formatand a DNA primer on a HER2 aptamer initiates auxiliary DNA self-assembled on the electrode to form a long one-dimensional DNA. The resulting DNA is then reacted with molybdate to generate electrochemical current. The sensitivity of the aptasensor with DNA self-assembly was greater than that of the aptasensor without DNA self-assembly due to the extended length of the DNA strand. Aptasensor analysis of HER2 in serum of breast cancer patients and healthy individuals is highly correlated (R2 = 0.9924) with ELISA measurements, with a p value of 1.37 × 10-7. The analysis of HER2 in serum (confirmed by ELISA) suggests that HER2 levels in breast cancer patients are much higher than healthy individuals. For HER2 positive patients, the levels are higher than those of HER2 negative patients. After surgery, there is a drop of HER2 levels in serum, suggesting potential clinical applications of the new self-assembled DNA electric current generating biosensor. Unlike proteins, DNA is easily amplifiable. The DNA signal amplification method presented here enables effective current generation, which can find wide range of biomedical applications for protein detection.

Immobilization and stability studies of multi-thiol probes for electrical biosensors by label-free analysis

ABSTRACTFor electrical biosensors based on gold electrodes, poor long-term stability of the monothiol-to-gold bonds has been a major issue for a robust and reliable performance of the sensor. The use of multi-thiol modified bioprobes was proposed to achieve a stable attachment. In this study, the binding kinetics and stability of mono/dithiol modified oligonucleotides were investigated using quartz crystal microbalance with dissipation (QCM-D) and electrochemical impedance spectroscopy (EIS). These label-free surface tools compared the binding, stability and the effect of dithiothreitol in solution of monothiol, dithiol, or disulfide modification. The potential for development of label-free, sensitive DNA microarrays is demonstrated.

Autonomous electrochemical biosensors: A new vision to direct methanol fuel cells

A new approach to biosensing devices is demonstrated aiming an easier and simpler application in routine health care systems. Our methodology considered a new concept for the biosensor transducing event that allows to obtain, simultaneously, an equipment-free, user-friendly, cheap electrical biosensor. The use of the anode triple-phase boundary (TPB) layer of a passive direct methanol fuel cell (DMFC) as biosensor transducer is herein proposed. For that, the ionomer present in the anode catalytic layer of the DMFC is partially replaced by an ionomer with molecular recognition capability working as the biorecognition element of the biosensor. In this approach, fuel cell anode catalysts are modified with a molecularly imprinted polymer (plastic antibody) capable of protein recognition (ferritin is used as model protein), inserted in a suitable membrane electrode assembly (MEA) and tested, as initial proof-of-concept, in a non-passive fuel cell operation environment. The anchoring of the ionomer-based plastic antibody on the catalyst surface follows a simple one-step grafting from approach through radical polymerization. Such modification increases fuel cell performance due to the proton conductivity and macroporosity characteristics of the polymer on the TPB. Finally, the response and selectivity of the bioreceptor inside the fuel cell showed a clear and selective signal from the biosensor. Moreover, such pioneering transducing approach allowed amplification of the electrochemical response and increased biosensor sensitivity by 2 orders of magnitude when compared to a 3-electrodes configuration system.

A review on ZnO-based electrical biosensors for cardiac biomarker detection.

Over the past few decades zinc oxide (ZnO)-based thin films and nanostructures have shown unprecedented performance in a wide range of applications. In particular, owing to high isoelectric point, biocompatibility and other multifunctional characteristics, ZnO has extensively been studied as a transduction material for biosensor development. The fascinating properties of ZnO help retain biological activity of the immobilized biomolecule and help in achieving enhanced sensing performance. As a consequence of recent advancements in this multidisciplinary field, diagnostic biosensors are expanding beyond traditional clinical labs to point-of-care and home settings. Label-free electrical detection of biomarkers has been demonstrated using ZnO-sensing platforms. In this review we highlight the characteristics of ZnO that enable realization of its use in development of point-of-care biosensors toward disease diagnosis, in particular cardiovascular diseases.

Silicon nitride surfaces as active substrate for electrical DNA biosensors

Miniaturization and integration are fundamental steps to fabricate smart and easy-to-use DNA biosensors to be massively exploited. Among the several materials, silicon nitride, extensively used as passivation layer in the microelectronics industry, is a good candidate for the development of biosensors employing electrical transduction. We report an exhaustive study on chemical grafting of DNA oligonucleotides on Si3N4 surfaces and electrical testing for DNA detection. In particular, we developed an optimised grafting protocol on Si3N4 surfaces characterised by contact angle, ellipsometry, Atomic Force Microscopy, and Transmission Electron Microscopy. A comparison with thermally grown SiO2 surfaces chemically treated with the same immobilization protocol was also carried out. DNA size electrical detection was carried out by immobilizing oligonucleotide probes on MIS (Metal Insulator Semiconductor)-like capacitors and measuring the characteristic capacitance voltage (C-V) curves upon hybridization with both oligo-perfect match (DNA PM) and PCR product targets. The correlation between DNA size and C-V shift has been evaluated. The analytical performances at various PM target concentrations showed a sensitivity value of 0.048V/nM with a LoD (Limit of Detection) of about bout 2.0nM and LoQ (Limit of Quantification) of 6.5nM. For PCR product target, the measured sensitivity was 0.043V/ngml−1 with a LoD of about 2.7ng/μl and LoQ of 8.8ng/μl.

(Invited) T-Chip, a Biosensor Based on the Electron Tunneling: Theory and Experiment

The presentation covers an electrical biosensor platform (T-chip) based on the electron tunneling phenomenon between the metal electrode and the methylene blue (MB) molecules. Firstly, the general idea of the T-chip is introduced. Secondly, the practical application of the T-chip for detecting biomarkers of the cancer is suggested. Finally, the numerical simulation of the T-chip considering the electron tunneling and the transport of ions in the electrolyte is introduced. To detect the target biomarkers in aqueous solution, the T-chip is integrated with the peptide molecules as the probe molecule of which end terminal was functionalized with the redox sites providing the electron states for tunneling. An asymmetric gold electrode structure of the T-chip facilitates a simple 2-electrode structure and reliable electrical measurement without adopting a reference electrode used in the conventional electrochemical system. As an example of the biosensor application, we selected the matrix metalloproteinase-9 (MMP-9) as the target biomarker which indicates metastasis of the cancer and tumor invasion. As the probe molecules, a peptide sequence conjugated with the MB was designed to be cleaved by the MMP-9. As an electrical indicator of the probe-target cleavage event, the MB was chosen. The cleavage event causes the reduction of the number of the MB sites (electron tunneling states) which leads a decrease in the electron tunneling current. To describe the physical model and evaluate the electrical characteristics of the T-chip, a numerical simulator (3-dimension) has been developed. To estimate the electron tunneling rates between the electrodes and the MB states, we used the non-equilibrium Green’s function (NEGF) method. Additionally, the ion motions in electrolyte is calculated by using drift-diffusion as the continuity equation. Finally, the terminal current due to the electron tunneling and ion movement was extracted by using the extended Ramo-Shockley theorem. Theory and experiment of the tunneling based biosensor will enhance the potential of success of the electrical biosensor through detailed understanding of the molecules and surface chemistry involved in the sensor system.