Characterisation of capacitive field-effect sensors with a nanocrystalline-diamond film as transducer material for multi-parameter sensing

The feasibility of a capacitive field-effect EDIS (electrolyte-diamond-insulator-semiconductor) platform for multi-parameter sensing is demonstrated by realising EDIS sensors with an O-terminated nanocrystalline-diamond (NCD) film as transducer material for the detection of pH and penicillin concentration as well as for the label-free electrical monitoring of adsorption and binding of charged macromolecules, like polyelectrolytes. The NCD films were grown on p-Si-SiO(2) substrates by microwave plasma-enhanced chemical vapour deposition. To obtain O-terminated surfaces, the NCD films were treated in an oxidising medium. The NCD-based field-effect sensors have been characterised by means of constant-capacitance method. The average pH sensitivity of the O-terminated NCD film was 40 mV/pH. A low detection limit of 5 microM and a high penicillin G sensitivity of 65-70 mV/decade has been obtained for an EDIS penicillin biosensor with the adsorptively immobilised enzyme penicillinase. Alternating potential changes, having tendency to decrease with increasing the number of adsorbed polyelectrolyte layers, have been observed after the layer-by-layer deposition of polyelectrolyte multilayers, using positively charged PAH (poly (allylamine hydrochloride)) and a negatively charged PSS (poly (sodium 4-styrene sulfonate)) as a model system. The response mechanism of the developed EDIS sensors is discussed.     

Progress of new label-free techniques for biosensors: a review

The detection techniques used in biosensors can be broadly classified into label-based and label-free. Label-based detection relies on the specific properties of labels for detecting a particular target. In contrast, label-free detection is suitable for the target molecules that are not labeled or the screening of analytes which are not easy to tag. Also, more types of label-free biosensors have emerged with developments in biotechnology. The latest developed techniques in label-free biosensors, such as field-effect transistors-based biosensors including carbon nanotube field-effect transistor biosensors, graphene field-effect transistor biosensors and silicon nanowire field-effect transistor biosensors, magnetoelastic biosensors, optical-based biosensors, surface stress-based biosensors and other type of biosensors based on the nanotechnology are discussed. The sensing principles, configurations, sensing performance, applications, advantages and restriction of different label-free based biosensors are considered and discussed in this review. Most concepts included in this survey could certainly be applied to the development of this kind of biosensor in the future.     

Detection of DNA and proteins using amorphous silicon ion-sensitive thin-film field effect transistors

Amorphous silicon-based ion-sensitive field-effect transistors (a-Si:H ISFETs) are used for the label-free detection of biological molecules. The covalent immobilization of DNA, followed by DNA hybridization, and of the surface adsorption of oligonucleotides and proteins were detected electronically by the a-Si:H ISFET. The ISFET measurements are performed with an external Ag/AgCl microreference electrode immersed in 100mM phosphate buffer electrolyte with pH 7.0. Threshold voltage shifts in the transfer curve of the ISFETs are observed resulting from successive steps of surface chemical functionalization, covalent DNA attachment to the functionalized surface, surface blocking, and hybridization with a complementary target. The surface sensitivity achieved for DNA oligonucleotides is of the order of 1pmol/cm(2). Point-of-zero charge estimations were made for the functionalized surfaces and for the device surface after DNA immobilization and hybridization. The results show a correlation between the changes in the point-of-zero charge and the shift observed in the threshold voltage of the devices. Electronic detection of adsorbed proteins and DNA is also achieved by monitoring the shifts of the threshold voltage of the ISFETs, with a sensitivity of approximately 50nM.     

Label-free detection of single nucleotide polymorphisms utilizing the differential transfer function of field-effect transistors

We present a label-free method for the detection of DNA hybridization, which is monitored by non-metallized silicon field-effect transistors (FET) in a microarray approach. The described method enables a fast and fully electronic readout of ex situ binding assays. The label-free detection utilizing the field-effect is based on the intrinsic charge of the DNA molecules and/or on changes of the solid–liquid interface impedance, when biomolecules bind to the sensor surface. With our sensor system, usually a time-resolved, dc readout is used. In general, this FET signal suffers from sensor drift, temperature drift, changes in electrolyte composition or pH value, influence of the reference electrode, etc. In this article, we present a differential ac readout concept for FET microarrays, which enables a stable operation of the sensor against many of these side-parameters, reliable readout and a possibility for a quick screening of large sensor arrays. We present the detection of point mutations in short DNA samples with this method in an ex situ binding assay.     

An enhanced glucose biosensor using charge transfer techniques

An enhanced glucose biosensor based on a charge transfer technique glucose sensor (CTTGS) is described and demonstrated experimentally. In the proposed CTTGS, which is accumulation method (d-gluconate+H(+)) ion perception system, the quality of output signal with "signal integration cycles" is high. With the proposed CTTGS it is possible to amplify the sensing signals without an external amplifier by using an accumulation cycle. It can be supposed that measurements of small (d-gluconate+H(+)) ion fluctuation are difficult by ion-sensitive field effect transistor (ISFET) because the theoretical maximum sensitivity is only 59 mV/pH and the small output signals are buried in the 1/f noise component of the metal-insulator-semi-conductor field-effect transistor (MISFET). Therefore, the CTTGS has many advantages, such as high sensitivity, high accuracy, high signal-to-noise ratio (SNR), and has been successfully demonstrated using a charge transfer technique. The CTTGS exhibited excellent performance for glucose with a large span (1445 mV) and good reproducibility. Moreover, the CTTGS has good sensitivity in this range of 7.22mV/mM, a lower detection limit of about 0.01 mM/L and an upper detection limit of about 200 mM/L compared with amperometric glucose analysis which has been studied recently. Under optimum conditions, the proposed CTTGS exceeds the performance of the widely used ISFET glucose sensor. The sensitivity of the CTTGS (7.22 mV/mM) was seven times higher than that of the ISFET (1 mV/mM). Furthermore, the sensitivity obtained for human glucose levels was 29.06 mV/mM with a non-linear error of +/-0.27%; the linearity is y=0.0294x+1.8612 and R(2)=0.9999, which is acceptable for clinical application. Real sample analysis is investigated in blood glucose level by our developed CTTGS ISFET system.     

Real-time and noninvasive monitoring of respiration activity of fertilized ova using semiconductor-based biosensing devices

In this report, we propose a novel evaluation method of embryo activity, describing the real-time and noninvasive electrical monitoring of embryo activity, caused by fertilization of the sea urchin, using a biologically-coupled field-effect transistor (bio-FET) comprised of semiconductor-based biosensing devices. The detection principle of bio-FET is based on the potentiometric detection of charge density change at the gate insulator, which includes changes of hydrogen ion concentration corresponding to pH variation. The surface potential at the gate surface of the bio-FET increased after the introduction of sperms into the ova, resulting in fertilization on the gate sensing area. The positive shift of surface potential indicates the increase of positive charges of hydrogen ions generated by dissolved carbon dioxide in artificial sea water based on respiration activity of the embryo. Moreover, the electrical signal of embryo activity is suppressed due to the inhibition of cytokinesis by introduction of cytochalasin B. The platform based on the bio-FET is expected to be a real-time, label-free and noninvasive detection system, not only in fundamental studies of embryo activity but also in the evaluation of embryo quality for in vitro fertilization.     

Functionalized SnO₂ nanobelt field-effect transistor sensors for label-free detection of cardiac troponin

Real-time label-free electrical detection of proteins, including cardiac troponin (cTn), is demonstrated using functionalized SnO? nanobelt field-effect transistors (FETs) with integrated microfluidics. Selective biomolecular functionalization of the active SnO? nanobelt channel and effective passivation of the substrate surface were realized and verified through fluorescence microscopy. The validation/verification of the sensing scheme was initially demonstrated via detection of biotin-streptavidin binding: devices with single biotinylated SnO? nanobelts showed pronounced conductance changes in response to streptavidin binding. Importantly, the pH-dependence of the conductance changes was fully consistent with the charged states of streptavidin at different pH. Moreover, the specificity of the sensors' electrical responses was confirmed by co-labeling with quantum dots. Finally, the sensing platform was successfully applied for detection of the cardiac troponin I (cTnI) subunit within cTn, a clinically important protein marker for myocardial infarction.     

Extended-gate FET-based enzyme sensor with ferrocenyl-alkanethiol modified gold sensing electrode

We developed a field-effect transistor (FET)-based enzyme sensor that detects an enzyme-catalyzed redox-reaction event as an interfacial potential change on an 11-ferrocenyl-1-undecanethiol (11-FUT) modified gold electrode. While the sensitivity of ion-sensitive FET (ISFET)-based enzyme sensors that detect an enzyme-catalyzed reaction as a local pH change are strongly affected by the buffer conditions such as pH and buffer capacity, the sensitivity of the proposed FET-based enzyme sensor is not affected by them in principle. The FET-based enzyme sensor consists of a detection part, which is an extended-gate FET sensor with an 11-FUT immobilized gold electrode, and an enzyme reaction part. The FET sensor detected the redox reaction of hexacyanoferrate ions, which are standard redox reagents of an enzymatic assay in blood tests, as a change in the interfacial potential of the 11-FUT modified gold electrode in accordance with the Nernstian response at a slope of 59 mV/decade at 25 degrees C. Also, the FET sensor had a dynamic range of more than five orders and showed no sensitivity to pH. A FET-based enzyme sensor for measuring cholesterol level was constructed by adding an enzyme reaction part, which contained cholesterol dehydrogenase and hexacyanoferrate (II)/(III) ions, on the 11-FUT modified gold electrode. Since the sensitivity of the FET sensor based on potentiometric detection was independent of the sample volume, the sample volume was easily reduced to 2.5 microL while maintaining the sensitivity. The FET-based enzyme sensor successfully detected a serum cholesterol level from 33 to 233 mg/dL at the Nernstian slope of 57 mV/decade.     

Non-Faradaic electrochemical detection of protein interactions by integrated neuromorphic CMOS sensors

Electronic detection of the binding event between biotinylated bovine serum albumen (BSA) and streptavidin is demonstrated with the chemoreceptive neuron MOS (CnuMOS) device. Differing from the ion-sensitive field-effect transistors (ISFET), CnuMOS, with the potential of the extended floating gate determined by both the sensing and control gates in a neuromorphic style, can provide protein detection without requiring analyte reference electrodes. In comparison with the microelectrode arrays, measurements are gathered through purely capacitive, non-Faradaic interactions across insulating interfaces. By using a (3-glycidoxypropyl)trimethoxysilane (3-GPS) self-assembled monolayer (SAM) as a simple covalent link for attaching proteins to a silicon dioxide sensing surface, a fully integrated, electrochemical detection platform is realized for protein interactions through monotone large-signal measurements or small-signal impedance spectroscopy. Calibration curves were created to coordinate the sensor response with ellipsometric measurements taken on witness samples. By monitoring the film thickness of streptavidin capture, a sensitivity of 25ng/cm2 or 2A of film thickness was demonstrated. With an improved noise floor the sensor can detect down to 2ng/(cm2mV) based on the calibration curve. AC measurements are shown to significantly reduce long-term sensor drift. Finally, a noise analysis of electrochemical data indicates 1/f(alpha) behavior with a noise floor beginning at approximately 1Hz.     

Silicon photonic crystal nanocavity-coupled waveguides for error-corrected optical biosensing

A photonic crystal (PhC) waveguide based optical biosensor capable of label-free and error-corrected sensing was investigated in this study. The detection principle of the biosensor involved shifts in the resonant mode wavelength of nanocavities coupled to the silicon PhC waveguide due to changes in ambient refractive index. The optical characteristics of the nanocavity structure were predicted by FDTD theoretical methods. The device was fabricated using standard nanolithography and reactive-ion-etching techniques. Experimental results showed that the structure had a refractive index sensitivity of 10(-2) RIU. The biosensing capability of the nanocavity sensor was tested by detecting human IgG molecules. The device sensitivity was found to be 2.3±0.24×10(5) nm/M with an achievable lowest detection limit of 1.5 fg for human IgG molecules. Additionally, experimental results demonstrated that the PhC devices were specific in IgG detection and provided concentration-dependent responses consistent with Langmuir behavior. The PhC devices manifest outstanding potential as microscale label-free error-correcting sensors, and may have future utility as ultrasensitive multiplex devices.     

Comparison of label-free biosensing in microplate, microfluidic, and spot-based affinity capture assays

Using both experimental assays and fluid-dynamic finite element simulation models, we directly compared the achievable performance limits of four distinct assay configurations for label-free detection of an analyte from a test sample on a biosensor surface. The assay configurations studied in this work included a biosensor incorporated into the bottom surface of a microplate well and a microfluidic channel. For each configuration, we compared assay performance for the scenario in which the entire bottom surface of the fluid-handling vessel is coated with capture ligands with assay performance for the scenario in which the capture ligands are applied in the form of localized spots. As a model system, we used detection of the protein biomarker tumor necrosis factor-alpha (TNF-α) using immobilized TNF-α capture antibody. Results show that the microfluidic assay format dramatically reduces the time required to establish a stable equilibrium. Spot-based assays are advantageous for microplate-based detection for reducing the time required for equilibrium sensor response. The results derived are generally applicable to any label-free biosensor technology and any ligand-analyte system with adjustable variables that include sensor mass density sensitivity, analyte-ligand adsorption/desorption rate constants, immobilized ligand density, flow channel geometry, flow rate, and spot size.     

Development of immunosensors for direct detection of three wound infection biomarkers at point of care using electrochemical impedance spectroscopy

A method for label-free, electrochemical impedance immunosensing for the detection and quantification of three infection biomarkers in both buffer and directly in the defined model matrix of mock wound fluid is demonstrated. Triggering Receptor-1 Expressed on Myeloid cells (TREM-1) and Matrix MetalloPeptidase 9 (MMP-9) are detected via direct assay and N-3-oxo-dodecanoyl-l-HomoSerineLactone (HSL), relevant in bacterial quorum sensing, is detected using a competition assay. Detection is performed with gold screen-printed electrodes modified with a specific thiolated antibody. Detection is achieved in less than 1h straight from mock wound fluid without any extensive sample preparation steps. The limits of detection of 3.3 pM for TREM-1, 1.1 nM for MMP-9 and 1.4 nM for HSL are either near or below the threshold required to indicate infection. A relatively large dynamic range for sensor response is also found, consistent with interaction between neighbouring antibody-antigen complexes in the close-packed surface layer. Together, these three novel electrochemical immunosensors demonstrate viable multi-parameter sensing with the required sensitivity for rapid wound infection detection directly from a clinically relevant specimen.     

An array of field-effect nanoplate SOI capacitors for (bio-)chemical sensing

An array of individually addressable nanoplate field-effect capacitive (bio-)chemical sensors based on an SOI (silicon-on-insulator) structure has been developed. The isolation of the individual capacitors was achieved by forming a trench in the top Si layer with a thickness of 350 nm. The realized sensor array allows addressable biasing and electrical readout of multiple nanoplate EISOI (electrolyte-insulator-silicon-on-insulator) capacitive biosensors on the same SOI chip as well as differential-mode measurements. The feasibility of the proposed approach has been demonstrated by realizing sensors for the pH and penicillin concentration detection as well as for the label-free electrical monitoring of polyelectrolyte multilayers formation and DNA (deoxyribonucleic acid)-hybridization event. A potential change of ～ 120 mV has been registered after the DNA hybridization for the sensor immobilized with perfectly matched single-strand DNA, while practically no signal changes have been observed for a sensor with fully mismatched DNA. The realized examples demonstrate the potential of the nanoplate SOI capacitors as a new basic structural element for the development of different types of field-effect biosensors.     

Electrical detection of VEGFs for cancer diagnoses using anti-vascular endotherial growth factor aptamer-modified Si nanowire FETs

We report the real-time, label-free and electrical detection of vascular endotherial growth factor (VEGF) for cancer diagnosis using anti-VEGF aptamer-modified Si nanowire field-effect transistors (SiNW-FETs). Specifically, the high quality single-crystalline SiNWs of both n-type and p-type characters were surface modified with the covalent immobilization of anti-VEGF aptamers, and they were turned into SiNW-FET biosensors for the VEGF detection. We show that the VEGF molecules consistently act on the gate dielectrics of both n-type and p-type SiNW-FETs as electrically positive point-charges; their recognition to anti-VEGF aptamers depletes (accumulates) the charge carriers in the p-type (n-type) SiNW-FETs and thus decreases (increases) the detection currents. The detection limit for VEGFs in this study was determined as 1.04nM and 104pM for the cases of n-type and p-type SiNW-FETs, respectively.     

Formaldehyde-sensitive sensor based on recombinant formaldehyde dehydrogenase using capacitance versus voltage measurements

A new formaldehyde-selective biosensor was constructed using NAD⁺- and glutathione-dependent recombinant formaldehyde dehydrogenase as a bio-recognition element immobilised on the surface of Si/SiO₂/Si₃N₄ structure. Sensor's response to formaldehyde was evaluated by capacitance measurements. The calibration curves obtained for formaldehyde concentration range from 10 μM to 20 mM showed a broad linear response with a sensitivity of 31 mV/decade and a detection limit about 10 μM. It has been shown that the output signal decreases with the increase of borate buffer concentration and the best sensitivity is observed in 2.5 mM borate buffer, pH 8.40. The response of the created formaldehyde-sensitive biosensor has also been examined in 2.5 mM Tris–HCl buffer, and the shift to the positive bias of the C(V) curves along with the potential axis has been observed, but the sensitivity of the biosensor in this buffer is decreased dramatically to the value of 2.4 mV/decade.     

A label-free immunosensor for determination of salbutamol based on localized surface plasmon resonance biosensing

We developed a localized surface plasmon resonance (LSPR)-based label-free optical biosensor for detection of salbutamol (Sal). Hollow gold nanoparticles (HGNs) which deposited on transparent indium tin oxide (ITO) film coated glass was used to sensing platform. Antibody against Sal was immobilized on HGN surface to recognize the target Sal molecules. Thus, the change of LSPR peak was proportional to the concentration of Sal in the solution. The experimental results demonstrated that the LSPR immunosensor possessed a good sensitivity and a high selectivity for Sal. The detection range for Sal was from 0.05 to 0.8 μg/mL with a correlation coefficient of 0.996. The biosensor was applied for the detection for Sal in spiked animal feed and pork liver samples, and the recoveries were in the range of 97–105 %. Therefore, it is expected that this approach may offer a new method in designing label-free LSPR immunosensor for detection of small molecules.     

Urea sensor with single poly(propylene) membrane

Urease was immobilized on the plasma-aminated surface of a hyfrophobic poly(propylene) (PP) membrane. This membrane, with urease matrix on one side while maintaining its original hydrophobic property on the other, was used to construct the urea sensor. The new urea sensors had response sensitivities ranged from 19 mV/decade to 30 mV/decade depending on the conditions of the plasma reaction. The enzyme electrode using single membrane gave a shorter response time as compared to the corresponding conventional electrode employing two seperate PP membranes. The sensitivity of the enzyme electrode increased with increasing buffer pH and reached a maximal level (40 mV/decade) at pH 7.6. The response sensitivity of the electrode was not affected by the change of buffer strength. Deamination of the plasma-modified hydrophobic PP membrane did not occur in aqueous environment judging from the stability of the urea electrode up to 12 days of operation. (c) 1992 John Wiley & Sons. Inc.     

3D antibody immobilization on a planar matrix surface

The amount of active capture antibodies immobilized per unit square is crucial to developing effective antibody chips, biosensors, immunoassays and other molecular recognition technologies. In this study, we present a novel yet simple method for oriented antibody immobilization at high density based on the formation of an orderly, organized aggregation of immunoglobulin G (IgG) and staphylococcal protein A (SPA). Following the chelation of His-tag with Ni(2+), antibodies were immobilized on a solid surface in a three-dimensional (3D) manner and exposed the analyte receptor sites well, which resulted in a substantial enhancement of the analytical signal with more than 64-fold increase in detection sensitivity. Pull-down assays confirmed that IgG antibody can only bind to Ni(2+) matrix indirectly via a SPA linkage, where the His-tag is responsible for binding Ni(2+) and homologous domains are responsible for binding IgG Fc. The immobilization approach proposed here may be an attractive strategy for the construction of high performance antibody arrays and biosensors as long as the antibody probe is of mammalian IgG.     

Rapid and sensitive biosensor for Salmonella

The rapid and sensitive detection of Salmonella typhymurium based on the use of a polyclonal antibody immobilized by the Langmuir-Blodgett method on the surface of a quartz crystal acoustic wave device was demonstrated. The binding of bacteria to the surface changed the crystal resonance parameters; these were quantified by the output voltage of the sensor instrumentation. The sensor had a lower detection limit of a few hundred cells/ml, and a response time of < 100 s over the range of 10(2)-10(10) cells/ml. The sensor response was linear between bacterial concentrations of 10(2)-10(7) cells/ml, with a sensitivity of 18 mV/decade. The binding of bacteria was specific with two binding sites needed to bind a single cell. The sensors preserve approximately 75% of their sensitivity over a period of 32 days.