Fabrication and characterization of microfabricated on-chip microelectrochemical cell for biosensing applications

The fabrication of on-chip microelectrochemical cell on Si wafer by means of photolithography is described here. The single on-chip microelectrochemical cell device has dimensions of 100 × 380 mm with integrated Pt counter electrode (CE), Ag/AgCl reference electrode (RE) and gold microelectrode array of 500 nm recess depth as the working electrode (WE). Two geometries of electrode array were implemented, band and disc, with fixed diameter/width of 10 µm; and varied centre-to-centre spacing (d) and number of electrodes (N) in the array. The on-chip microelectrochemical cell structure has been designed to facilitate further WE biomodifications. Firstly, the developed microelectrochemical cell does not require packaging hence reducing the production cost and time. Secondly, the working electrode (WE) on the microelectrochemical cell is positioned towards the end of the chip enabling modification of the working electrode surface to be carried out for surface bio-functionalisation without affecting both the RE and CE surface conditions. The developed on-chip microelectrochemical cell was examined with scanning electron microscopy (SEM) and characterised by two electrochemical techniques. Both cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were performed in 1 mM ferrocenecarboxylic acid (FCA) in 0.01 M phosphate buffered saline (PBS) solution at pH7.4. Electrochemical experiments showed that in the case of halving the interspacing distance of the microdisc WE array (50 nm instead of 100 nm), the voltammogram shifted from a steady-state CV (feature of hemispherical diffusion) to an inclined peak-shaped CV (feature of linear diffusion) albeit the arrays had the same surface area. In terms of EIS it was also found that linear diffusion dominates the surface instead of hemispherical diffusion once the interspacing distance was reduced, supporting the fact that closely packed arrays may behave like a macroelectrode

System packaging & integration for a swallowable capsule using a direct access sensor

Technological developments in biomedical microsystems are opening up new opportunities to improve healthcare procedures. Swallowable diagnostic capsules are an example of this. In this paper, a diagnostic capsule technology is described based on direct-access sensing of the Gastro Intestinal (GI) fluids throughout the GI tract. The objective of this paper is two-fold: i) develop a packaging method for a direct access sensor, ii) develop an encapsulation method to protect the system electronics. The integrity of the interconnection after sensor packaging and encapsulation is correlated to its reliability and thus of importance. The zero level packaging of the sensor was achieved by using a so called Flip Chip Over Hole (FCOH) method. This allowed the fluidic sensing media to interface with the sensor, while the rest of the chip including the electrical connections can be insulated effectively. Initial tests using Anisotropic Conductive Adhesive (ACA) interconnect for the FCOH demonstrated good electrical connections and functionality of the sensor chip. Also a preliminary encapsulation trial of the flip chipped sensor on a flexible test substrate has been carried out and showed that silicone encapsulation of the system is a viable option

Chemically modified electrodes for recessed microelectrode array

Chemical modifications on recessed microelectrode array, achieved via electrodeposition techniques are reported here. Silicon-based gold microelectrode arrays of 10µm microband and microdisc array were selected and functionalised using sol-gel and nanoporous gold (NPG) respectively. For electrochemically assisted self-assembly (EASA) formati6154on of sol-gel, electrode surface was first pre-treated with a self-assembled partial monolayer of mercaptopropyltrimethoxysilane (MPTMS) before transferring it into the sol containing cetyltrimethyl ammonium bromide (CTAB)/tetraethoxysilane (TEOS):MPTMS (90:10) precursors. A cathodic potential is then applied. It was found that larger current densities were required in ensuring successful film deposition when moving from macro- to micro- dimensions. For NPG modification, a chemical etching process called dealloying was employed. NPG of three different thicknesses have been successfully deposited. All the modified and functionalized microelectrode arrays were characterized by both optical (SEM) and electrochemical analysis (cyclic voltammetry and impedance spectroscopy). An increase in surface area and roughness has been observed and such will benefit for future sensing application

Ultra-Sensitive determination of pesticides via cholinesterase-based sensors for environmental analysis.

This review is focussed towards the development of acetylcholinesterase enzymatic based biosensors for the quantification of trace concentrations of highly toxic pesticides via their inhibitory effect on the enzyme. Initial results were obtained using wild-type enzymes which have a broad spectrum of susceptibility to a variety of pesticides. The sensitivity and selectivity of the enzyme activity was improved by development and screening of a wide range of mutant enzymes. Optimal enzymes were then exploited within a range of sensor formats. A range of immobilisation techniques including adsorption based approaches, binding via proteins and entrapment within conducting polymers were all studied. The incorporation of stabilisers and co-factors were utilised to optimise electrode performance and stability - with both planar and microelectrode geometries being developed. Reproducible quantification of pesticides could be obtained at concentrations down to 10-17 M, representing a detection limit hitherto unavailable

System packaging & integration for a swallowable capsule using a direct access sensor

Technological developments in biomedical microsystems are opening up new opportunities to improve healthcare procedures. Swallowable diagnostic capsules are an example of this. In this paper, a diagnostic capsule technology is described based on direct-access sensing of the Gastro Intestinal (GI) fluids throughout the GI tract. The objective of this paper is two-fold: i) develop a packaging method for a direct access sensor, ii) develop an encapsulation method to protect the system electronics. The integrity of the interconnection after sensor packaging and encapsulation is correlated to its reliability and thus of importance. The zero level packaging of the sensor was achieved by using a so called Flip Chip Over Hole (FCOH) method. This allowed the fluidic sensing media to interface with the sensor, while the rest of the chip including the electrical connections can be insulated effectively. Initial tests using Anisotropic Conductive Adhesive (ACA) interconnect for the FCOH demonstrated good electrical connections and functionality of the sensor chip. Also a preliminary encapsulation trial of the flip chipped sensor on a flexible test substrate has been carried out and showed that silicone encapsulation of the system is a viable option

Nanoenabling electrochemical sensors for life sciences applications

Electrochemical sensing systems are advancing into a wide range of new applications, moving from the traditional lab environment into disposable devices and systems, enabling real-time continuous monitoring of complex media. This transition presents numerous challenges ranging from issues such as sensitivity and dynamic range, to autocalibration and antifouling, to enabling multiparameter analyte and biomarker detection from an array of nanosensors within a miniaturized form factor. New materials are required not only to address these challenges, but also to facilitate new manufacturing processes for integrated electrochemical systems. This paper examines the recent advances in the instrumentation, sensor architectures, and sensor materials in the context of developing the next generation of nanoenabled electrochemical sensors for life sciences applications, and identifies the most promising solutions based on selected well established application exemplars

Fabrication and characterization of a test platform integrating nanoporous structures with biochemical functionality

The application of solid-state nanopore technology for biosensing is a rapidly developing area of research with high commercial potential. Different synthetic materials, including silicon nitride, alumina, and polymers, are employed to fabricate single and multiple pores and offer a good platform for selective biomolecule detection. Two solid-state pore arrays, one with integrated silicon microfluidic system, were considered and an immobilization strategy suitable for detecting a single-stranded DNA (ssDNA) sequence was investigated. For the silicon nitride pores, a modification method based on the use of 3-aminopropyltriethoxysilane for silanization and 1,4-phenylene diisothiocyanate for amine crosslinking was applied to immobilize 100-nM ssDNA (amine C6) and a 100-nM limit of detection for complementary to probe ssDNA (Cy5) was estimated. The polycarbonate pores (the second type of the pore arrays) underwent surface modification based on an oxidation reduction reaction using sodium periodate and sodium borohydride and was used to immobilize 10-nM ssDNA and an estimated 100-nM limit of detection was also achieved. Linear sweep voltammetry was used to characterize the pores and a current potential profile was obtained after both immobilization of probe ssDNA and hybridization of complementary to probe ssDNA on the modified pore array surface. A decrease in current amplitude was measured after surface modification of both pore arrays, and this was attributed to the appearance of an additional layer on the pore surface reducing the pore opening and hindering the current flow. The hybridization event was also supported by contact angle measurements, where an increase in hydrophilicity was recorded at the different surface modification steps that were applied to produce the biofunctionalized nanopore. In addition, fluorescence was observed on the surfaces after hybridization, through incorporation of a CY5 fluorescent tag attached on the 5' end of the complementary to probe DNA. These results show the potential to use both silicon nitride and polycarbonate nanopores in DNA detection applications

The Determinants of Oxygen Gradients in Respiring Samples and their Impact on Cellular Responses to Hypoxia

Quenched-phosphorescence oxygen (O2) sensing technique allows non-invasive, real-time monitoring of both intra- and extracellular O2 concentration in respiring samples. Using this technique we investigated O2 gradients in populations of neurosecretory PC12 cells cultured in 96-well plates and exposed to graded hypoxia at rest and upon metabolic stimulation. Under high atmospheric O2 (10–21%) the respiration of resting cells dictated that local O2 was moderately reduced, and at a certain threshold (6% in galactose medium) cell layer became practically anoxic. Furthermore, cell stimulation triggered a major redistribution of O2 and a prominent ‘hypoxic overshoot’ mediated by diffusion. The deep, prolonged cell deoxygenation upon stimulation was matched by an increase in nuclear HIF-1α levels. In the presence of nitric oxide the hypoxic overshoot was truncated and HIF-1α stabilization inhibited. Thus, the main determinants which impact upon cellular O2 levels and oxygen-sensitive signaling pathways are the atmospheric O2, sample geometry, cell density, respiration rate and its dynamics. Changes in any of these parameters can significantly alter the O2 levels experienced by the cells and the subsequently activated signaling pathways. This technique, which provides simple and reliable monitoring of cell oxygenation, is therefore important for hypoxia research, metabolic studies and experiments with respiring cells.Science Foundation Ireland, grant 07/IN.1/B1804 and Enterprise Ireland, grant EI CFTD/07/124Deposited by bulk impor