Micro- and nano-devices for electrochemical sensing

Electrode miniaturization has profoundly revolutionized the field of electrochemical sensing, opening up unprecedented opportunities for probing biological events with a high spatial and temporal resolution, integrating electrochemical systems with microfluidics, and designing arrays for multiplexed sensing. Several technological issues posed by the desire for downsizing have been addressed so far, leading to micrometric and nanometric sensing systems with different degrees of maturity. However, there is still an endless margin for researchers to improve current strategies and cope with demanding sensing fields, such as lab-on-a-chip devices and multi-array sensors, brain chemistry, and cell monitoring. In this review, we present current trends in the design of micro-/nano-electrochemical sensors and cutting-edge applications reported in the last 10 years. Micro- and nanosensors are divided into four categories depending on the transduction mechanism, e.g., amperometric, impedimetric, potentiometric, and transistor-based, to best guide the reader through the different detection strategies and highlight major advancements as well as still unaddressed demands in electrochemical sensing

Design and Implementation of an Integrated Biosensor Platform for Lab-on-a-Chip Diabetic Care Systems

Recent advances in semiconductor processing and microfabrication techniques allow the implementation of complex microstructures in a single platform or lab on chip. These devices require fewer samples, allow lightweight implementation, and offer high sensitivities. However, the use of these microstructures place stringent performance constraints on sensor readout architecture. In glucose sensing for diabetic patients, portable handheld devices are common, and have demonstrated significant performance improvement over the last decade. Fluctuations in glucose levels with patient physiological conditions are highly unpredictable and glucose monitors often require complex control algorithms along with dynamic physiological data. Recent research has focused on long term implantation of the sensor system. Glucose sensors combined with sensor readout, insulin bolus control algorithm, and insulin infusion devices can function as an artificial pancreas. However, challenges remain in integrated glucose sensing which include degradation of electrode sensitivity at the microscale, integration of the electrodes with low power low noise readout electronics, and correlation of fluctuations in glucose levels with other physiological data. This work develops 1) a low power and compact glucose monitoring system and 2) a low power single chip solution for real time physiological feedback in an artificial pancreas system. First, glucose sensor sensitivity and robustness is improved using robust vertically aligned carbon nanofiber (VACNF) microelectrodes. Electrode architectures have been optimized, modeled and verified with physiologically relevant glucose levels. Second, novel potentiostat topologies based on a difference-differential common gate input pair transimpedance amplifier and low-power voltage controlled oscillators have been proposed, mathematically modeled and implemented in a 0.18μm [micrometer] complementary metal oxide semiconductor (CMOS) process. Potentiostat circuits are widely used as the readout electronics in enzymatic electrochemical sensors. The integrated potentiostat with VACNF microelectrodes achieves competitive performance at low power and requires reduced chip space. Third, a low power instrumentation solution consisting of a programmable charge amplifier, an analog feature extractor and a control algorithm has been proposed and implemented to enable continuous physiological data extraction of bowel sounds using a single chip. Abdominal sounds can aid correlation of meal events to glucose levels. The developed integrated sensing systems represent a significant advancement in artificial pancreas systems

Multisite monitoring of choline using biosensor microprobe arrays in combination with CMOS circuitry

A miniature device enabling parallel in vivo detection of the neurotransmitter choline in multiple brain regions of freely behaving rodents is presented. This is achieved by combining a biosensor microprobe array with a custom-developed CMOS chip. Each silicon microprobe comprises multiple platinum electrodes that are coated with an enzymatic membrane and a permselective layer for selective detection of choline. The biosensors, based on the principle of amperometric detection, exhibit a sensitivity of 157±35 µA mM-1 cm-2, a limit of detection of below 1 µM, and a response time in the range of 1 s. With on-chip digitalization and multiplexing, parallel recordings can be performed at a high signal-to-noise ratio with minimal space requirements and with substantial reduction of external signal interference. The layout of the integrated circuitry allows for versatile configuration of the current range and can, therefore, also be used for functionalization of the electrodes before use. The result is a compact, highly integrated system, very convenient for on-site measurement

Biosensor system with an integrated CMOS microelectrode array for high spatio-temporal electrochemical imaging, A

2019 Fall.Includes bibliographical references.The ability to view biological events in real time has contributed significantly to research in life sciences. While optical microscopy is important to observe anatomical and morphological changes, it is equally important to capture real-time two-dimensional (2D) chemical activities that drive the bio-sample behaviors. The existing chemical sensing methods (i.e. optical photoluminescence, magnetic resonance, and scanning electrochemical), are well-established and optimized for existing ex vivo or in vitro analyses. However, such methods also present various limitations in resolution, real-time performance, and costs. Electrochemical method has been advantageous to life sciences by supporting studies and discoveries in neurotransmitter signaling and metabolic activities in biological samples. In the meantime, the integration of Microelectrode Array (MEA) and Complementary-Metal-Oxide-Semiconductor (CMOS) technology to the electrochemical method provides biosensing capabilities with high spatial and temporal resolutions. This work discusses three related subtopics in this specific order: improvements to an electrochemical imaging system with 8,192 sensing points for neurotransmitter sensing; comprehensive design processes of an electrochemical imaging system with 16,064 sensing points based on the previous system; and the application of the system for imaging oxygen concentration gradients in metabolizing bovine oocytes. The first attempt of high spatial electrochemical imaging was based on an integrated CMOS microchip with 8,192 configurable Pt surface electrodes, on-chip potentiostat, on-chip control logic, and a microfluidic device designed to support ex vivo tissue experimentation. Using norepinephrine as a target analyte for proof of concept, the system is capable of differentiating concentrations of norepinephrine as low as 8µM and up to 1,024 µM with a linear response and a spatial resolution of 25.5×30.4μm. Electrochemical imaging was performed using murine adrenal tissue as a biological model and successfully showed caffeine-stimulated release of catecholamines from live slices of adrenal tissue with desired spatial and temporal resolutions. This system demonstrates the capability of an electrochemical imaging system capable of capturing changes in chemical gradients in live tissue slices. An enhanced system was designed and implemented in a CMOS microchip based on the previous generation. The enhanced CMOS microchip has an expanded sensing area of 3.6×3.6mm containing 16,064 Pt electrodes and the associated 16,064 integrated read channels. The novel three-electrode electrochemical sensor system designed at 27.5×27.5µm pitch enables spatially dense cellular level chemical gradient imaging. The noise level of the on-chip read channels allow amperometric linear detection of neurotransmitter (norepinephrine) concentrations from 4µM to 512µM with 4.7pA/µM sensitivity (R=0.98). Electrochemical response to dissolved oxygen concentration or oxygen partial pressure (pO2) was also characterized with deoxygenated deionized water containing 10µM to 165 µM pO2 with 8.21pA/µM sensitivity (R=0.89). The enhanced biosensor system also demonstrates selectivity to different target analytes using cyclic voltammetry to simultaneously detect NE and uric acid. In addition, a custom-designed indium tin oxide and Au glass electrode is integrated into the microfluidic support system to enable pH measurement, ensuring viability of bio-samples in ex vivo experiments. Electrochemical images confirm the spatiotemporal performance at four frames per second while maintaining the sensitivity to target analytes. The overall system is controlled and continuously monitored by a custom-designed user interface, which is optimized for real-time high spatiotemporal resolution chemical bioimaging. It is well known that physiological events related to oxygen concentration gradients provide valuable information to determine the state of metabolizing biological cells. Utilizing the CMOS microchip with 16,064 Pt MEA and an improved three-electrode system configuration, the system is capable of imaging low oxygen concentration with limit of detection of 18.3µM, 0.58mg/L, or 13.8mmHg. A modified microfluidic support system allows convenient bio-sample handling and delivery to the MEA surface for sensing. In vitro oxygen imaging experiments were performed using bovine cumulus-oocytes-complexes cells with custom software algorithms to analyze its flux density and oxygen consumption rate. The imaging results are processed and presented as 2D heatmaps, representing the dissolved oxygen concentration in the immediate proximity of the cell. The 2D images and analysis of oxygen consumption provide a unique insight into the spatial and temporal dynamics of cell metabolism

Multisite monitoring of choline using biosensor microprobe arrays in combination with CMOS circuitry

A miniature device enabling parallel in vivo detection of the neurotransmitter choline in multiple brain regions of freely behaving rodents is presented. This is achieved by combining a biosensor microprobe array with a custom-developed CMOS chip. Each silicon microprobe comprises multiple platinum electrodes that are coated with an enzymatic membrane and a permselective layer for selective detection of choline. The biosensors, based on the principle of amperometric detection, exhibit a sensitivity of 157 +/- 35 mu A mM(-1) cm(-2), a limit of detection of below 1 mu M, and a response time in the range of 1 s. With on-chip digitalization and multiplexing, parallel recordings can be performed at a high signal-to-noise ratio with minimal space requirements and with substantial reduction of external signal interference. The layout of the integrated circuitry allows for versatile configuration of the current range and can, therefore, also be used for functionalization of the electrodes before use. The result is a compact, highly integrated system, very convenient for on-site measurements

A fully integrated CMOS microelectrode system for electrochemistry

Electroanalysis has proven to be one of the most widely used technologies for point-of-care devices. Owing to the direct recording of the intrinsic properties of biochemical functions, the field has been involved in the study of biology since electrochemistry’s conception in the 1800’s. With the advent of microelectronics, humanity has welcomed self-monitoring portable devices such as the glucose sensor in its everyday routine. The sensitivity of amperometry/ voltammetry has been enhanced by the use of microelectrodes. Their arrangement into microelectrode arrays (MEAs) took a step forward into sensing biomarkers, DNA and pathogens on a multitude of sites. Integrating these devices and their operating circuits on CMOS monolithically miniaturised these systems even more, improved the noise response and achieved parallel data collection. Including microfluidics on this type of devices has led to the birth of the Lab-on-a-Chip technology. Despite the technology’s inclusion in many bioanalytical instruments there is still room for enhancing its capabilities and application possibilities. Even though research has been conducted on the selective preparation of microelectrodes with different materials in a CMOS MEA to sense several biomarkers, limited effort has been demonstrated on improving the parallel electroanalytical capabilities of these devices. Living and chemical materials have a tendency to alter their composition over time. Therefore analysing a biochemical sample using as many electroanalytical methods as possible simultaneously could offer a more complete diagnostic snapshot. This thesis describes the development of a CMOS Lab-on-a-Chip device comprised of many electrochemical cells, capable of performing simultaneous amperometric/voltammetric measurements in the same fluidic chamber. The chip is named an electrochemical cell microarray (ECM) and it contains a MEA controlled by independent integrated potentiostats. The key stages in this work were: to investigate techniques for the electrochemical cell isolation through simulations; to design and implement a CMOS ECM ASIC; to prepare the CMOS chip for use in an electrochemical environment and encapsulate it to work with liquids; to test and characterise the CMOS chip housed in an experimental system; and to make parallel measurements by applying different simultaneous electroanalytical methods. It is envisaged that results from the system could be combined with multivariate analysis to describe a molecular profile rather than only concentration levels. Simulations to determine the microelectrode structure and the potentiostat design, capable of constructing isolated electrochemical cells, were made using the Cadence CAD software package. The electrochemical environment and the microelectrode structure were modelled using a netlist of resistors and capacitors. The netlist was introduced in Cadence and it was simulated with potentiostat designs to produce 3-D potential distribution and electric field intensity maps of the chemical volume. The combination of a coaxial microelectrode structure and a fully differential potentiostat was found to result in independent electrochemical cells isolated from each other. A 4 x 4 integrated ECM controlled by on-chip fully differential potentiostats and made up by a 16 × 16 working electrode MEA (laid out with the coaxial structure) was designed in an unmodified 0.35 μm CMOS process. The working electrodes were connected to a circuit capable of multiplexing them along a voltammetric measurement, maintaining their diffusion layers during stand-by time. Two readout methods were integrated, a simple resistor for an analogue readout and a discrete time digital current-to-frequency charge-sensitive amplifier. Working electrodes were designed with a 20 μm side length while the counter and reference electrodes had an 11 μm width. The microelectrodes were designed using the aluminium top metal layer of the CMOS process. The chips were received from the foundry unmodified and passivated, thus they were post-process fabricated with photolithographic processes. The passivation layer had to be thinned over the MEA and completely removed on top of the microelectrodes. The openings were made 25 % smaller than the top metal layer electrode size to ensure a full coverage of the easily corroded Al metal. Two batches of chips were prepared, one with biocompatible Au on all the microelectrodes and one altered with Pd on the counter and Ag on the reference electrode. The chips were packaged on ceramic pin grid array packages and encapsulated using chemically resistant materials. Electroplating was verified to deposit Au with increased roughness on the microelectrodes and a cleaning step was performed prior to electrochemical experiments. An experimental setup containing a PCB, a PXIe system by National Instruments, and software programs coded for use with the ECM was prepared. The programs were prepared to conduct various voltammetric and amperometric methods as well as to analyse the results. The first batch of post-processed encapsulated chips was used for characterisation and experimental measurements. The on-chip potentiostat was verified to perform alike a commercial potentiostat, tested with microelectrode samples prepared to mimic the coaxial structure of the ECM. The on-chip potentiostat’s fully differential design achieved a high 5.2 V potential window range for a CMOS device. An experiment was also devised and a 12.3 % cell-to-cell electrochemical cross-talk was found. The system was characterised with a 150 kHz bandwidth enabling fast-scan cyclic voltammetry(CV) experiments to be performed. A relatively high 1.39 nA limit-of-detection was recorded compared to other CMOS MEAs, which is however adequate for possible applications of the ECM. Due to lack of a current polarity output the digital current readout was only eligible for amperometric measurements, thus the analogue readout was used for the rest of the measurements. The capability of the ECM system to perform independent parallel electroanalytical measurements was demonstrated with 3 different experimental techniques. The first one was a new voltammetric technique made possible by the ECM’s unique characteristics. The technique was named multiplexed cyclic voltammetry and it increased the acquisition speed of a voltammogram by a parallel potential scan on all the electrochemical cells. The second technique measured a chemical solution with 5 mM of ferrocene with constant potential amperometry, staircase cyclic voltammetry, normal pulse voltammetry, and differential pulse voltammetry simultaneously on different electrochemical cells. Lastly, a chemical solution with 2 analytes (ferrocene and decamethylferrocene) was prepared and they were sensed separately with constant potential amperometry and staircase cyclic voltammetry on different cells. The potential settings of each electrochemical cell were adjusted to detect its respective analyte

Towards Single Bacterium Detection: A Microelectronic/Microfluidic Hybrid System Based on a CMOS Technology

RÉSUMÉ Cette thèse porte sur le développement d'un biocapteur hybride CMOS microfluidique capable de détecter des bactéries pathogènes une à une en temps réel basé sur un principe de spectroscope impédimétrique. Le biocapteur proposé se compose d'une matrice de capteurs qui comportent une matrice de microélectrodes, desmultiplexeurs à commande numérique, et des circuits de détection intégrés sur une puce de silicium CMOS. Cette recherche propose une nouvelle structure de microélectrodes qui permet à une structure de microélectrodes face à face à haute densité intégrable par post-traitement d’une puce CMOS. Au lieu d’être créée par le dépôt et la gravure de couches métalliques supplémentaires, la structure de microélectrodes face à face est construite en exploitant un empilement de couches métalliques disponible avec la technologie CMOS adoptée. Les détecteurs sont obtenus en construisant des microcanaux qui traversent le substrat. Ces microcanaux passent entre les microélectrodes face à face. Lorsque les fluides où se trouvent les échantillons traversent le microcanal, le système détecte de façon continue les changements d'impédance entre les microélectrodes induits par le passage de chaque bactérie . Cette thèse étudie le processus de microfabrication qui permet de libérer la matrice de microélectrodes et de fabriquer les microcanaux traversant le substrat. Les techniques dites de FIB (pours Focused Ion Beam) et de DRIE (pour Deep Reactive Ion Etching) sont utilisées. Les forces et faiblesses de chaque technologie sont analysées et des recettes de processus optimisés sont étudiées. La matrice de microélectrodes a été réalisée avec succès par les deux technologies. Comme preuve de concept, plusieurs microcanaux traversant le substrat sont également formés en utilisant la technologie FIB. Cette thèse propose également un nouveau circuit de détection. Réalisé grâce à la micro-électronique, ce circuit est capable de détecter les changements d'impédance causés par le passage d’une seule bactérie dans un milieu conducteur. Sans conditionnement de signaux et de circuit de traitement complexes, tels que des amplificateurs de haute précision, des filtres ou des convertisseurs analogue à numérique ou numérique à analogique, les circuits de détection sont conçus pour offrir une bonne sensibilité et une configurabilité qui permet de l'adapter aux différentes conditions de détection. Une technique de mise en boîtier biocompatible est également mise en oeuvre pour encapsuler le capteur intégré tout en fournissant des interfaces fluidiques et électriques pour l'injection d'échantillons et de signaux électriques. Une nouvelle approche pour améliorer la sélectivité de détection basée sur l’utilisation de bactéries magnétotactiques est également proposée dans cette thèse. Sous le contrôle d’un champ magnétique extérieur, les bactéries magnétotactiques sont utilisées comme bio-transporteurs, qui peuvent chercher activement et capturer les bactéries pathogènes cibles afin de les amener à la zone de détection. Une puce microfluidique est fabriquée grâce à des techniques de prototypage rapide afin de valider les idées proposées et de fournir des guides de conception d'une puce plus avancés. Les résultats de microfabrication et les résultats des tests préliminaires montrent que l'intégration monolithique des technologies CMOS et microfluidique est possible et qu’elle permet la réalisation de microélectrodes face à face dans une plate-forme capable de détecter le passage d’une seule bactérie en isolation.----------ABSTRACT This thesis reports on the development of a CMOS Microfluidic hybrid biosensor technology that is proposed to detect single pathogenic bacterium in real time based on impedimetric spectroscopy. The proposed biosensor consists of a CMOS silicon die that incorporates a microelectrode array, digitally controlled multiplexers, and sensing circuits. This research proposes a novel microelectrode structure, which is obtained by first manufacturing high density face to face microelectrodes on a CMOS die, possible by a relatively simple CMOS post-processing. Instead of deposition and patterning of additional metal layers, the face to face microelectrode array is constructed by stacking metal and via layers of the adopted CMOS technology. By constructing through substrate microchannels in between pairs of face to face microelectrodes, when a fluid sample flows through the microchannel, the microelectrodes on the wall detect the impedance change induced by bacterium in the fluid in a continuous way. This thesis investigates the microfabrication process of releasing microelectrode arrays and constructing through substrate microchannels. FIB (Focused Ion Beam) and DRIE (Deep Reactive Ion Etching) technologies are utilized. The strength and weakness of each technology are analyzed and optimized process recipes are investigated. Microelectrode array were successfully released using both process technologies. As a proof of concept, several through substrate microchannels were also formed by using the FIB technology. This thesis also proposes a novel sensing microelectronic circuit, which is able to sense the impedance change caused by a single bacterium in a conductive medium. The system does not require complex signal conditioning and processing circuits, such as high precision amplifiers, filters or ADC/DAC. The proposed simple sensing structure offer high sensitivity, reliability and configurability. A dedicated biocompatible packaging is also implemented to encapsulate the CMOS die and provide a microchamber, fluidic and electrical interfaces for sample injection and signal interfaces. A new approach to achieve detection selectivity or specificity assisted by magnetotactic bacterium is also proposed in this thesis. Under the control of an external magnetic field, the viii magnetotactic bacteria are used as bio-carriers, which can actively search and capture some target pathogenic bacteria and bring them to the sensing area. A microfluidic chip is fabricated by rapid prototyping techniques to validate the proposed idea and to provide design guides for a more advanced and highly integrated CMOS chip. The achieved microfabrication results and preliminary testing results show that the monolithic integration of CMOS and microfluidic technology, especially the face to face microelectrode structure is a suitable platform for single bacterium detection and analysis

Review on carbon-derived, solid-state, micro and nano sensors for electrochemical sensing applications

The aim of this review is to summarize the most relevant contributions in the development of electrochemical sensors based on carbon materials in the recent years. There have been increasing numbers of reports on the first application of carbon derived materials for the preparation of an electrochemical sensor. These include carbon nanotubes, diamond like carbon films and diamond film-based sensors demonstrating that the particular structure of these carbon material and their unique properties make them a very attractive material for the design of electrochemical biosensors and gas sensors. Carbon nanotubes (CNT) have become one of the most extensively studied nanostructures because of their unique properties. CNT can enhance the electrochemical reactivity of important biomolecules and can promote the electron-transfer reactions of proteins (including those where the redox center is embedded deep within the glycoprotein shell). In addition to enhanced electrochemical reactivity, CNT-modified electrodes have been shown useful to be coated with biomolecules (e.g., nucleic acids) and to alleviate surface fouling effects (such as those involved in the NADH oxidation process). The remarkable sensitivity of CNT conductivity with the surface adsorbates permits the use of CNT as highly sensitive nanoscale sensors. These properties make CNT extremely attractive for a wide range of electrochemical sensors ranging from amperometric enzyme electrodes to DNA hybridization biosensors. Recently, a CNT sensor based fast diagnosis method using non-treated blood assay has been developed for specific detection of hepatitis B virus (HBV) (human liver diseases, such as chronic hepatitis, cirrhosis, and hepatocellular carcinoma caused by hepatitis B virus). The linear detection limits for HBV plasma is in the range 0.5–3.0 μL−1 and for anti- HBVs 0.035–0.242 mg/mL in a 0.1 M NH4H2PO4 electrolyte solution. These detection limits enables early detection of HBV infection in suspected serum samples. Therefore, non-treated blood serum can be directly applied for real-time sensitive detection in medical diagnosis as well as in direct in vivo monitoring. Synthetic diamond has been recognized as an extremely attractive material for both (bio-) chemical sensing and as an interface to biological systems. Synthetic diamond have outstanding electrochemical properties, superior chemical inertness and biocompatibility. Recent advances in the synthesis of highly conducting nanocrystalline-diamond thin films and nano wires have lead to an entirely new class of electrochemical biosensors and bio-inorganic interfaces. In addition, it also combines with development of new chemical approaches to covalently attach biomolecules on the diamond surface also contributed to the advancement of diamond-based biosensors. The feasibility of a capacitive field-effect EDIS (electrolyte-diamond-insulatorsemiconductor) platform for multi-parameter sensing is demonstrated with an O-terminated nanocrystalline-diamond (NCD) film as transducer material for the detection of pH and penicillin concentration. This has also been extended for the label-free electrical monitoring of adsorption and binding of charged macromolecules. One more recent study demonstrated a novel bio-sensing platform, which is introduced by combination of a) geometrically controlled DNA bonding using vertically aligned diamond nano-wires and b) the superior electrochemical sensing properties of diamond as transducer material. Diamond nanowires can be a new approach towards next generation electrochemical gene sensor platforms. This review highlights the advantages of these carbon materials to promote different electron transfer reactions specially those related to biomolecules. Different strategies have been applied for constructing carbon material-based electrochemical sensors, their analytical performance and future prospects are discussed