Eric Moore

Comparison of Cell-Based Biosensors with Traditional Analytical Techniques for Cytotoxicity Monitoring and Screening of Polycyclic Aromatic Hydrocarbons in the Environment

Abstract Today, special attention is paid to the environmental pollution effects of polycyclic aromatic hydrocarbons (PAHs), which are carcinogenic and mutagenic compounds. They are also widely distributed in the environment and can be found in water, soil, and plants. For this reason, various technologies have been developed to detect and screen for PAHs, and one of the recent developments is cell-based biosensors (CBBs). This type of sensor allows for real-time measurements by detecting the impedance of cellular behavior, which has been proven sensitive to toxic compounds. This articles presents a review of the previous work on CBBs for environmental monitoring of PAHs, in comparison with other conventional methods, which are based on chromatographic techniques and immunoassay.

Development of a biosensor for the quantitative detection of 2,4,6-trichloroanisole using screen printed electrodes

Immunoassay techniques can provide a simple, practical and inexpensive method for analysis of 2,4,6-trichloroanisole (TCA). Enzyme linked immunosorbent assay (ELISA) and electrochemical techniques were investigated with the purpose of attaining high selectivity and sensitivity. Both assays incorporated a direct format for analysis of TCA using alkaline phosphatase (AP), as the labeling species. TCA has no functional groups through which linkage of enzymes or proteins can be achieved so alternative hapten molecules were developed. Molecular recognition between TCA and the antibodies that were raised against haptens whose chemical structures were similar to the target analyte was good. This was especially true for the competition between free analyte and hapten A-AP conjugate for the protein G purified polyclonal antibodies (pAb 76). ELISA results using the direct format of analysis were poor when compared to the achieved results for the electrochemical sensor. A limit of detection of 1 ng/ml (1 ppb) was achieved for the best ELISA system while a limit of detection of 29 pg/ml (29 ppt) was obtained for the electrochemical sensor.

Development of Polyclonal Antibodies Against Domoic Acid for Their Use in Electrochemical Biosensors

Abstract The current work describes the raising of polyclonal antibodies (pAbs) against domoic acid (DA), an algal toxin produced by the diatom Pseudonitzschia pungens. They were screened for sensitivity and selectivity using a competitive enzyme-linked immunosorbent assay (ELISA). The antiserum produced against a keyhole limpet haemocyanin (DA-KLH) conjugate displayed a high affinity for free DA. The optimized horseradish peroxidase (HRP) ELISA had a detection limit of 0.6 ng mL−1 (ppb) and a working range of 0.8–300 ppb DA applying a streptavidin-biotin amplification system (ABC system). Furthermore this antiserum did not cross-react with similar chemical structures and algal toxins such as kainic acid, aspartic acid, glutamic acid, geranic acid and 2-methyl-3-butenoic acid. When the ELISA was compared using an alkaline phosphatase (AP) label we found the system to behave in a similar manner to the optimized HRP system but the linear range was smaller in the high DA concentration range. These pAbs were then used in the optimization of a screen-printed electrode (SPE) system for measurement of DA. A disposable screen-printed carbon electrode coupled with amperometric detection of p-aminophenol at +300 mV vs. Ag/AgCl, produced by the enzyme AP, was used for signal measurement. The sensor incorporates a relevant range for toxin detection, by which humans become ill (Iverson, F.; Truelove, J. Toxicology and seafood toxins: domoic acid. Natural Toxins 1994, 2, 334–339.) with detection limits achieved at SPE to the order of ppb. The SPE system is simple and cost-effective due to its disposable nature, and analysis time is complete in 30 min. In addition, recovery experiments on DA for both ELISA and SPE highlighted the functionality of these systems yielding a ±12% deviation for the true value for the ELISA using AP and ±25% for the sensor.

Electrochemical Immunosensors for Food Analysis: A Review of Recent Developments

ABSTRACT Electrochemical immunosensors have the potential to transform analytical procedures within the food industry by providing highly specific, rapid, and inexpensive determination of pathogens. In this paper, recent advances in this area are outlined. In particular, attention is paid to new methods that have been developed for the modification of working electrode surfaces. Many advances have been related to the use of novel nanomaterials such as carbon nanotubes, graphene, and metallic nanoparticles, often used in conjunction with each other or polymers. The use of these materials has generally provided superior sensor sensitivity. The application of immunosensors to the detection of a range of pathogens in real samples is then investigated to establish whether they provide solutions in practical applications.

In vitro fibroblast and pre-osteoblastic cellular responses on laser surface modified Ti-6Al-4V.

The success of any implant, dental or orthopaedic, is driven by the interaction of implant material with the surrounding tissue. In this context, the nature of the implant surface plays a direct role in determining the long term stability as physico-chemical properties of the surface affect cellular attachment, expression of proteins, and finally osseointegration. Thus to enhance the degree of integration of the implant into the host tissue, various surface modification techniques are employed. In this work, laser surface melting of titanium alloy Ti-6Al-4V was carried out using a CO2 laser with an argon gas atmosphere. Investigations were carried out to study the influence of laser surface modification on the biocompatibility of Ti-6Al-4V alloy implant material. Surface roughness, microhardness, and phase development were recorded. Initial knowledge of these effects on biocompatibility was gained from examination of the response of fibroblast cell lines, which was followed by examination of the response of osteoblast cell lines which is relevant to the applications of this material in bone repair. Biocompatibility with these cell lines was analysed via Resazurin cell viability assay, DNA cell attachment assay, and alamarBlue metabolic activity assay. Laser treated surfaces were found to preferentially promote cell attachment, higher levels of proliferation, and enhanced bioactivity when compared to untreated control samples. These results demonstrate the tremendous potential of this laser surface melting treatment to significantly improve the biocompatibility of titanium implants in vivo.

An Integrated Optofluidic Platform for DNA Hybridization and Detection

There has been extensive research into micro total analysis systems (micro-TAS) and lab-on-a-chip research due to the benefits of increased sample throughput, reduced sample consumption, and rapid analysis times. The integration of low-cost fluidic and optical components offers the possibility of complex systems with increased functionality on a single detection platform. For the development of an integrated optofluidic system for DNA hybridization, the key areas are optical/fluidic integration and the efficiency of surface chemistry integration within the system. The impact of fluidic parameters such as flow rate, channel height, and time on hybridization performance is to optimize detection performance over conventional assay (microtiter plate formats). The use of a passive waveguide device means DNA binding events can be monitored using fluorescence excitation or refractive index measurement. The integration of the three areas is enhanced by the robustness of the waveguide material (oxide, nitride), enabling chemical functionalization by initial silanization followed by addition of a linker molecule 1,4 phenylene diisothiocyanate (PDITC) for covalent immobilization of DNA probes together with the possibility to define microfluidics on the waveguide substrate using standard SU8 photolithography. The fluidic design requires 240 nl of analyte to fill the integrated optofluidic system. Here, we report the novel integration and optimization of a covalent surface chemistry with microfluidic channels for fluidic delivery, and a standard resonant mirror (RM) waveguide detection platform. The optofluidic detection platform was tested using fluorescence and refractive index to monitor binding events between target and probe DNA. We describe the detection system, using simulations to explain the response to changes in refractive index and outline a method for covalent attachment of DNA probes surface chemistry protocol to immobilize probe DNA on the sensor surface and the optimization of fluidic design, achieving pM detection limit. We highlight the benefit of optimizing the fluidic component and its benefit in hybridization efficiency an approach often overlooked in sensor design and performance.

Comparative Study of 4-Aminophenyl Phosphate and Ascorbic Acid 2-Phosphate, as Substrates for Alkaline Phosphatase Based Amperometric Immunosensor

Abstract The performance of ascorbic acid 2-phosphate (AAP), an alternative substrate for alkaline phosphatase (AP), was compared with a widely used 4-aminophenyl phosphate (pAPP). AP is used here as a label in enzyme immunoassay with electrochemical detection. Linear sweep voltammetry and amperometry were used. Optimal working potentials were investigated at four electrode materials such as Pt, Au, glassy carbon, and carbon screen printed electrode (SPE). Two configurations of SPE were investigated, such as amperometry in a stirred batch and a single-drop analysis at a coplanar three-electrode strip. In both systems, at all four electrodes, pAP required significantly lower detection potentials yielding significantly higher signals. Enzyme kinetics of both substrates with AP was examined. The system has been applied to an enzyme immunoassay of rabbit IgG with AP as a label.

Amperometric Immunosensors for screening of Polycyclic Aromatic Hydrocarbons in water

An amperometric immunosensor with low limit detection was developed for the screening of polycyclic aromatic hydrocarbons (PAHs) in water. The system was based on detecting the specific substance using an immunological reaction by measuring the chemical responses to specific antibodies. An integrated biochip with a three electrode system was fabricated. Gold was used as the working electrode with platinum was used as the counter electrode. A modified Ag/AgCl reference electrode was employed to enhance the stability of the immunosensors. Indirect competition enzyme-linked immunosorbent assay (ELISA) was carried out within the electrode using alkaline phosphatase (AP) as the labelled-enzyme. The system shows acceptable reproducibility and good stability. The immunosensor exhibited a wide linear response to PAHs. A limit of detection for this sensor was in the range of 1 to 10 ng ml−1 in aqueous sample.

Integration of a 3D hydrogel matrix within a hollow core photonic crystal fibre for DNA probe immobilization

In this paper, we demonstrate the integration of a 3D hydrogel matrix within a hollow core photonic crystal fibre (HC-PCF). In addition, we also show the fluorescence of Cy5-labelled DNA molecules immobilized within the hydrogel formed in two different types of HC-PCF. The 3D hydrogel matrix is designed to bind with the amino groups of biomolecules using an appropriate cross-linker, providing higher sensitivity and selectivity than the standard 2D coverage, enabling a greater number of probe molecules to be available per unit area. The HC-PCFs, on the other hand, can be designed to maximize the capture of fluorescence to improve sensitivity and provide longer interaction lengths. This could enable the development of fibre-based point-of-care and remote systems, where the enhanced sensitivity would relax the constraints placed on sources and detectors. In this paper, we will discuss the formation of such polyethylene glycol diacrylate (PEGDA) hydrogels within a HC-PCF, including their optical properties such as light propagation and auto-fluorescence.

Nerve localization techniques for peripheral nerve block and possible future directions

Ultrasound guidance is now a standard nerve localization technique for peripheral nerve block (PNB). Ultrasonography allows simultaneous visualization of the target nerve, needle, local anesthetic injectate, and surrounding anatomical structures. Accurate deposition of local anesthetic next to the nerve is essential to the success of the nerve block procedure. Due to limitations in the visibility of both needle tip and nerve surface, the precise relationship between needle tip and target nerve is unknown at the moment of injection. Importantly, nerve injury may result both from an inappropriately placed needle tip and inappropriately placed local anesthetic. The relationship between the block needle tip and target nerve is of paramount importance to the safe conduct of peripheral nerve block.