Advancements in the micro-and nano-scale fabrication techniques have opened up new avenues for the development of portable, scalable and easier-to-use biosensors. Over the last few years, electrodes made of carbon have been widely used as sensing units in biosensors due to their attractive physiochemical properties. The aim of this research is to investigate different strategies to develop functionalized high surface carbon micro/nano-structures for electrochemical and biosensing devices. High aspect ratio three-dimensional carbon microarrays were fabricated via carbon microelectromechanical systems (C-MEMS) technique, which is based on pyrolyzing pre-patterned organic photoresist polymers. To further increase the surface area of the carbon microstructures, surface porosity was introduced by two strategies, i.e. (i) using F127 as porogen and (ii) oxygen reactive ion etch (RIE) treatment. Electrochemical characterization showed that porous carbon thin film electrodes prepared by using F127 as porogen had an effective surface area (Aeff 185%) compared to the conventional carbon electrode. To achieve enhanced electrochemical sensitivity for C-MEMS based functional devices, graphene was conformally coated onto high aspect ratio three-dimensional (3D) carbon micropillar arrays using electrostatic spray deposition (ESD) technique. The amperometric response of graphene/carbon micropillar electrode arrays exhibited higher electrochemical activity, improved charge transfer and a linear response towards H2O2 detection between 250μM to 5.5mM. Furthermore, carbon structures with dimensions from 50 nano-to micrometer level have been fabricated by pyrolyzing photo-nanoimprint lithography patterned organic resist polymer. Microstructure, elemental composition and resistivity characterization of the carbon nanostructures produced by this process were very similar to conventional photoresist derived carbon. Surface functionalization of the carbon nanostructures was performed using direct amination technique. Considering the need for requisite functional groups to covalently attach bioreceptors on the carbon surface for biomolecule detection, different oxidation techniques were compared to study the types of carbon–oxygen groups formed on the surface and their percentages with respect to different oxidation pretreatment times. Finally, a label-free detection strategy using signaling aptamer/protein binding complex for platelet-derived growth factor oncoprotein detection on functionalized three-dimensional carbon microarrays platform was demonstrated. The sensor showed near linear relationship between the relative fluorescence difference and protein concentration even in the sub-nanomolar range with an excellent detection limit of 5 pmol.

Carbon fiber ultramicroelectrodes (CF-UMEs) are commonly used as electrochemical probes and sensors due to their small size, fast response, and high signal-to-noise ratio. Surface modification strategies are often employed on CF-UMEs to improve their selectivity and sensitivity for desired applications. However, many modification methods are cumbersome and require expensive equipment. In this study, a simple approach known as soft nitriding is used to prepare nitrogen-doped CF-UMEs (N-CF-UMEs). Nitrogen groups introduced via soft nitriding act as electrocatalytic sites for the breakage of O-O bonds during the reduction of peroxides like H2O2, a common target of biosensing strategies. Voltammetric studies confirm that, compared to CF-UMEs, N-CF-UMEs possess enhanced electrocatalytic activity towards H2O2 reduction as evidenced by an increase in current and positive shift in onset potential for the reaction. N-CF-UMEs also proved capable for amperometric detection of H2O2, exhibiting good linear response from 0.1 to 5.6 mM at -0.4 V vs. Ag/AgCl.

Early and accessible diagnostics are important elements to reducing the negative side-effects of untreated disease. One key advancement in diagnostic monitoring is through the development of highly sensitive sensors that have the capability to detect lower concentrations, while still remaining accessible for point-of-care use. This dissertation characterizes electrochemical sensing platforms that are enhanced by convection and by 3D electrodes made from high surface area, vertically aligned carbon nanotubes (VACNTs). Free-standing VACNTs were patterned into microchannel arrays for flow-through amperometric sensing. Convective mass transfer enhancement was shown to improve sensor performance in amperometric sensing through the use of high surface area to fluid volume structures and concentration boundary layer confinement. Through-flow sensing of hydrogen peroxide produced drastically higher signals than stirred sensing, with over 90% of the hydrogen peroxide being oxidized as it passed through the channels. Non-enzymatic sensing of glucose was achieved by chemical reaction of glucose with methyl viologen to produce on average 3.4 electrons per glucose molecule, significantly higher than that obtained with enzymatic sensing with glucose oxidase. A scaled down sensor enabled detection from 200 μL of glucose by flow injection analysis with a limit of detection of 360 nM and a linear sensing range up to at least 150 μM glucose. Such sensing range offers the potential to measure glucose levels found in saliva. This work demonstrates the utility of high aspect ratio electrodes made of VACNTs. Convection and surface area are shown to enhance the sensitivity of flow-through VACNT amperometric sensors by effectively utilizing the available analyte to increase the measured current density. Advances in nanomaterials, combined with electrochemical impedance spectroscopy, have allowed impedimetric biosensors to have high sensitivity while remaining label-free, pushing towards enabling portable diagnosis at the point-of-care. Porous, 3D VACNT electrodes for impedance-based biosensing were fabricated with different electrode height, gap width, and configuration. Sensitivity was characterized by functionalizing the representative protein streptavidin onto VACNT electrodes for detection of biotin. Tall, closely-spaced VACNT interdigitated electrodes are shown to have the highest electroactive surface area (15x the 2D geometric area) and the highest sensitivity, allowing for a 1 ng/mL limit of detection. Aspect ratio and surface area are shown to be important factors in determining the sensitivity of 3D VACNT interdigitated electrodes for impedimetric sensing of biomolecules bound to electrode surfaces. Although this biosensing platform is shown with streptavidin and biotin, it could be extended to other proteins, antibodies, viruses, and bacteria.

Compared to unmodified carbons and even some metal materials, nitrogen-doped carbons have been found to exhibit better performance for reducing oxygen-oxygen bonds, a key step in electroreduction of both O2 (an important reaction in energy applications) and H2O2 (an important reaction in sensing and biosensing). Previous studies from our lab revealed that thermal decomposition of urea in the presence of carbon fiber (CF) results in N-doped that exhibited good electrocatalytic properties for H2O2 reduction. However, previous methods of sealing ultramicroelectrodes (UMEs) made from N-doped CF using laser heating of borosilicate capillaries and epoxy seemed to affect surface nitrogen contents and electrocatalytic properties. In this work, we evaluate paraffin sealing as a strategy for preparing UMEs in a way that minimizes effects on important surface nitrogen species so that electrocatalytic properties of the N-doped CF towards H2O2 reduction can be retained.

Urine culture, the current gold standard for urinary tract infection (UTI) diagnosis, does not produce results in an acceptable length of time. An ultra-sensitive, cost-effective electrochemical biosensing platform with nanostructured microelectrodes was designed to address the need for a rapid, point-of-care (PoC) test that could achieve a sample-to-answer time in less than an hour. Printed circuit boards and metallized glass slides were processed using various techniques and then tested for their ability to form nanostructured microelectrodes. Peptide nucleic acid probes for the bacteria and yeast as well as ten probes for antibiotic resistance genes were designed and synthesized for use with the new platform. Validation of the sensor's specificity was performed using high concentrations (100nM) of synthetic DNA oligomers. Furthermore, a clinically relevant sensitivity of 103 cfu/mL was demonstrated by detecting 4 pathogen lysates (Staphylococcus saprophyticus, Pseudomonas aeruginosa, Enterococcus faecalis and Klebsiella pneumoniae) in a buffered solution.

Urine culture, the current gold standard for urinary tract infection (UTI) diagnosis, does not produce results in an acceptable length of time. An ultra-sensitive, cost-effective electrochemical biosensing platform with nanostructured microelectrodes was designed to address the need for a rapid, point-of-care (PoC) test that could achieve a sample-to-answer time in less than an hour. Printed circuit boards and metallized glass slides were processed using various techniques and then tested for their ability to form nanostructured microelectrodes. Peptide nucleic acid probes for the bacteria and yeast as well as ten probes for antibiotic resistance genes were designed and synthesized for use with the new platform. Validation of the sensor's specificity was performed using high concentrations (100nM) of synthetic DNA oligomers. Furthermore, a clinically relevant sensitivity of 103 cfu/mL was demonstrated by detecting 4 pathogen lysates (Staphylococcus saprophyticus, Pseudomonas aeruginosa, Enterococcus faecalis and Klebsiella pneumoniae) in a buffered solution.

The rapid development of micro/nanofabrication techniques have enabled engineering of material interfacial properties. Micro/nanostructures with unique electrical, mechanical, thermal, magnetic, optical, and biological properties, have found applications in a wide range of fields such as electronics, photonics, biological/chemical sensing, tissue engineering, and diagnostics, etc. As such, numerous strategies have been developed for structuring materials into micro/nano- scale. However, the challenge still lies in the high cost, low throughput, complexity in fabrication, and difficulty in scaling up. This thesis aims to explore fabrication strategies for micro/nanostructured surfaces that are versatile, simple, and inexpensive. The thin film stencil lift-off technique with both Parylene and self-adhesive vinyl has been explored for this purpose. Further applications of the resulted micro/nanostructured surfaces are also presented in this thesis. Through improved Parylene stencil fabrication process, both spontaneously phase-segregated and arbitrary binary supported lipid bilayer patterns have been achieved. Also, the microstructured Parylene surfaces have been ddemonstrated for patterning stacked SLBs that are either homogeneous or phase-segregated. Without any lithography technique involved, vinyl stencil lift-off offers as a facile and inexpensive benchtop method for patterning thin films such as metal and glassy films. Combining the thermal shrinking of shape memory polymer, the patterned feature sizes are further decreased by 60% in both x and y dimensions, pushing the patterning resolution to down to sub-100 μm range. In addition, the shrinking process induces micro/nanostructures onto the deposited thin film, and the structure sizes are easily tunable with film thickness deposited. Further applications of such patterned micro/nanostructured surfaces has also been explored. The structured gold films have served as high-surface-area electrodes for electrochemical sensing. By introducing photoresist as a sacrificial layer, the structured gold thin films can be lifted off and transferred onto elastomeric substrate, and serve stretchable and flexible sensors. Such sensing devices exhibit great stability and reproducibility even when working under external strain. Finally, the micro/nanostructured glassy surfaces have been employed as substrate for cell growth to study topographical effect on cell morphology. It has been concluded that rougher surfaces lead to cell elongation, and finer structures promote filopodia generation. These results underscore the strength and suitability of thin film stencil lift-off as a powerful technique for creating micro- and nanostructured surfaces. These structured surfaces could find applications in many other areas, due to their great properties such as tunable structure size, high surface area, flexibility, and long-term stability. / Thesis / Doctor of Philosophy (PhD)

Dans cette thèse, nous présentons la détection électrochimique des herbicides, c'est-à-dire le paraquat et l'isoproturon dans des échantillons aqueux. Leur utilisation intensive est une source de contamination de l'environnement et leur toxicité constitue une menace pour la santé. La détection électrochimique est une technique prometteuse et avantageuse par rapport aux méthodes de détection conventionnelles en raison de ses propriétés telles que l'analyse rapide, la facilité d'utilisation, la rentabilité et la sensibilité élevée résultant de la modification de l'électrode de travail. Ici, nous avons modifié les électrodes modifiées avec des films minces de silice mésoporeuse pour agir comme capteurs d'herbicide. Ces électrodes ont été modifiées par un processus d'auto-assemblage assisté par électrochimie, un processus bien établi pour la modification des électrodes par notre groupe. Dans la première partie, l'adhérence du film de silice mésoporeux aux électrodes de carbone a été améliorée à l'aide d'une amine primaire qui a agi comme colle moléculaire pour une meilleure fixation de ces films à la surface des électrodes. Dans la partie suivante, ces électrodes modifiées ont été utilisées pour la détection électrochimique des herbicides susmentionnés. Les électrodes modifiées ont montré une sensibilité accrue et une limite de détection basse par rapport aux électrodes non modifiées. L'effet des différents paramètres de la solution ainsi que l'épaisseur du film et la géométrie de l'électrode ont également été étudiés et ont un impact critique sur la sensibilité du système / In this thesis, we present the electrochemical detection of herbicides i.e. paraquat and isoproturon in aqueous samples. These herbicides are used worldwide extensively for weed control in different crops. Their intensive use is a source of environmental contamination and their toxicity is a threat to Human health. Electrochemical sensing is a promising and advantageous technique as compared to conventional detection methods due to its properties such as rapid analysis, ease of operation, cost effectiveness and high sensitivity as a result of working electrode modification. Here, we modified electrodes modified with mesoporous silica thin films to act as herbicide sensors. These electrodes were modified by electrochemically assisted self-assembly process, a well-established process for electrode modification by our group. In the first part adhesion of mesoporous silica film at carbon electrodes was improved with the help of a primary amine which acted as molecular glue for better attachment of these films at electrodes surface. In the next part these modified electrodes were used for electrochemical detection of above stated herbicides. Modified electrodes showed enhanced sensitivity and low limit of detection as compared to unmodified ones. Effect of different solution parameters as well as film thickness and electrode geometry was also studied and found to have critical impact on sensitivity of the system

D. Tech. (Department of Chemistry, Faculty of Applied and Computer Sciences), Vaal University of Technology. / In this study, few layer graphene (Gr) and heteroatom graphene (HGr) were synthesized by chemical vapour deposition (CVD) method. Acetylene gas was used as carbon source for the synthesis of graphene, while a mixture of nitrobenzene and dichloromethane (ratio 1:1) were used as both carbon and dopant sources for the synthesis of the heteroatom graphene (HGr). A mixture of argon and nitrogen gases were carefully combined and used as carrier gasses and purge for both the synthesis of graphene and the synthesis of heteroatom graphene. X-ray diffraction (XRD) characterized showed that the as synthesized materials were crystalline materials, Raman spectroscopy indicated that the synthesized materials consist of sp2 hybridized carbon atoms, while scanning electron microscopy (SEM) and atomic force microscopy (AFM) results showed that the synthesized materials possess regions of 2 to 7 nm of thickness. Transmission electron microscopy (TEM) characterization also showed that the synthesized heteroatom graphene possesses about 5 to 7 layers with about 2 nm thickness, and x-ray photoelectron spectroscopy (XPS) result showed the presence of nitrogen, oxygen and chlorine in the lattice of the synthesized heteroatom graphene while the synthesized material still retained about 80% sp2 hybridization. The synthesized materials were used in the fabrication of modified bioelectrodes for electrobiocatalytic biosensing of glucose and hydroquinone. The fabricated bioelectrodes were characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The CV characterization showed a diffusion-controlled electrode processes in al modified electrodes, while the EIS characterization showed the presence of both diffusion controlled and kinetic controlled impedance at the electrode-electrolyte interface. The fabricated GC/PEDOT-PSS/HGr/Lac modified bioelectrode exhibited a kinetic controlled impedance of 3150 Ω, while the fabricated GC/PEDOT-PSS/Gr/Lac modified bioelectrode exhibited a kinetic controlled impedance of 4138 Ω. Chronoamperometric experiments showed that the fabricated bioelectrodes exhibited swift electrobiocatalytic activity towards glucose and hydroquinone sensing respectively for graphene and heteroatom graphene. The graphene modified bioelectrode exhibited a linear response of 0.2 to 9.8 mM glucose concentration and a sensitivity of 87.0 μA/mM/cm2, while the heteroatom modified bioelectrode also exhibited a swift response to step by step addition of hydroquinone with a limit of detection of 2.07 μM and dynamic range of 2.07μM to 2.97 mM, thus indicating the tremendous potential of the materials in a wide range of electrobiocatalytic and bioelectronics applications.

Diese Arbeit präsentiert eine einfache eine einfache, auf Photolithographie basierende Methode zur Darstellung einer isolierten Graphenkante (oder GrEdge) einer Monolage als Nanoelektrode auf einem isolierenden Substrat vorgestellt. Trotz ihrer Millimeter-Länge verhält sich die nur einen Nanometer breite GrEdge-Elektrode wie ein Nanodraht mit einem hohen Seitenverhältnis von 1000000 zu 1. Des Weiteren wird der Einsatz von elektrochemischer Modifikation (ECM) demonstriert, um die GrEdge selektiv mit Metall-Nanopartikeln und organischen Schichten nicht-kovalente oder kovalente zu funktionalisieren, wodurch die Chemie der Kante verändert werden kann. Durch die Anbringung von Metall-Nanopartikeln kann zusätzlich oberflächenverstärkte Raman-Spektroskopie (SERS) genutzt werden, um die chemische Beschaffenheit sowohl der unberührten als auch der funktionalisierten GrEdge zu charakterisieren. Die GrEdge weist sehr hohe Mass-entransportraten auf, was charakteristisch für Nanoelektroden ist. Dementsprechend wird die voltammetrische Antwort von der Kinetik des heterogenen Elektrontransfers (HET) diktiert. An der GrEdge-Elektrode werden hohe HET-Raten beobachtet: mindestens 14 cm/s für Außensphäre sonde Ferrocenmethanol (FcMeOH) mit einem quasi-Nernst'schen Verhalten und 0,06 cm/s oder höher für innere Sphäre sonde Ferricyanide ([Fe(CN)6]3-) mit einer kinetisch kontrollierten Reaktion. Nach der selektiven Modifikation der Kante mit Goldnanopartikeln erweist sich der HET als reversibel, mit einer massentransportbegrenztes Nernst‘sches Verhalten aufweisen für beide Redoxmoleküle. Darüber hinaus ermöglicht die schnelle HET-Kinetik die Detektion der reduzierten Form von Nicotinamid-Adenin-Dinukleotid (NADH) und Flavin-Adenin-Dinukleotid (FAD) mit niedrigen Ansatzpotentialen und hinunter bis zu niedrigen mikromolaren Konzentrationen. Entsprechend verbessert die vorliegende Arbeit das Verständnis der Kante von Graphen und deren Chemie. / This thesis presents a simple photolithography-based method to realize the isolated monolayer graphene edge (or GrEdge) nanoelectrode on an insulating substrate. The millimeter-long and a nanometer-wide GrEdge is found to behave like a nanowire with a high aspect ratio of 1000000-to-1. Further, the use of electrochemical modification (ECM) is demonstrated to selectively functionalize the GrEdge with metal nanoparticles and organic moieties in a non-covalent/ covalent manner to tune the chemistry of the edge. The attachment of metal nanoparticles was used to exploit surface-enhanced Raman scattering (SERS) to characterize the chemistry of both the pristine and the functionalized GrEdge. The GrEdge electrodes were found to exhibit very high mass transport rates, characteristic of nanoelectrodes. Accordingly, the voltammetric response is found to be dictated by the kinetics of heterogeneous electron transfer (HET), attributed to the nanoscale geometry and a unique diffusional profile at such electrodes. At the GrEdge electrode, high HET rates are observed: at least 14 cm/s for outer-sphere probe, ferrocenemethanol (FcMeOH) with a quasi-Nernstian behavior; and 0.06 cm/s or higher for inner-sphere probe, ferricyanide ([Fe(CN)6]3-) with a kinetically controlled response. Upon selective modification of the edge with gold nanoparticles, the HET is found to be reversible, with a mass-transport-limited Nernstian response for both probes. Furthermore, the fast HET kinetics enables the sensing of the reduced form of nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) with low onset potentials and down to low micromolar concentrations. Hence, this thesis improves the understanding of the edges of graphene and their chemistry. It also realizes isolated GrEdge as a new class of nanoelectrode which forms an important basis within the fields of fundamental electrochemistry and analytical sciences.