Synthetic Receptors for Early Detection and Treatment of Cancer.

Over recent decades, synthetic macrocyclic compounds have attracted interest from the scientific community due to their ability to selectively and reversibly form complexes with a huge variety of guest moieties. These molecules have been studied within a wide range of sensing and other fields. Within this review, we will give an overview of the most common synthetic macrocyclic compounds including cyclodextrins, calixarenes, calixresorcinarenes, pillarenes and cucurbiturils. These species all display the ability to form a wide range of complexes. This makes these compounds suitable in the field of cancer detection since they can bind to either cancer cell surfaces or indeed to marker compounds for a wide variety of cancers. The formation of such complexes allows sensitive and selective detection and quantification of such guests. Many of these compounds also show potential for the detection and encapsulation of environmental carcinogens. Furthermore, many anti-cancer drugs, although effective in in vitro tests, are not suitable for use directly for cancer treatment due to low solubility, inherent instability in in vivo environments or an inability to be adsorbed by or transported to the required sites for treatment. The reversible encapsulation of these species in a macrocyclic compound can greatly improve their solubility, stability and transport to required sites where they can be released for maximum therapeutic effect. Within this review, we intend to present the use of these species both in cancer sensing and treatment. The various macrocyclic compound families will be described, along with brief descriptions of their synthesis and properties, with an outline of their use in cancer detection and usage as therapeutic agents. Their use in the sensing of environmental carcinogens as well as their potential utilisation in the clean-up of some of these species will also be discussed.

Characterisation of thin films of graphene-surfactant composites produced through a novel semi-automated method.

In this paper we detail a novel semi-automated method for the production of graphene by sonochemical exfoliation of graphite in the presence of ionic surfactants, e.g., sodium dodecyl sulfate (SDS) and cetyltrimethylammonium bromide (CTAB). The formation of individual graphene flakes was confirmed by Raman spectroscopy, while the interaction of graphene with surfactants was proven by NMR spectroscopy. The resulting graphene-surfactant composite material formed a stable suspension in water and some organic solvents, such as chloroform. Graphene thin films were then produced using Langmuir-Blodgett (LB) or electrostatic layer-by-layer (LbL) deposition techniques. The composition and morphology of the films produced was studied with SEM/EDX and AFM. The best results in terms of adhesion and surface coverage were achieved using LbL deposition of graphene(-)SDS alternated with polyethyleneimine (PEI). The optical study of graphene thin films deposited on different substrates was carried out using UV-vis absorption spectroscopy and spectroscopic ellipsometry. A particular focus was on studying graphene layers deposited on gold-coated glass using a method of total internal reflection ellipsometry (TIRE) which revealed the enhancement of the surface plasmon resonance in thin gold films by depositing graphene layers.

Enhancement of electrode performance by a simple casting method using sonochemically exfoliated graphene.

We demonstrate within this paper a method for modifying commercial screen-printed electrodes with aqueous graphene suspensions to enhance their electrochemical activity. The graphene suspensions are synthesized by a simple ultrasonic exfoliation method from graphite, where reaggregation is prevented by the addition of common cationic or anionic surfactants, thereby avoiding the use of organic solvents or harsh chemical procedures. These suspensions can then be simply cast onto the screen-printed electrodes. Cyclic voltammetry with a number of redox active species such as phenols, as well as impedance measurements, were made to characterize these systems. The modified electrodes are shown to demonstrate significantly enhanced electrochemical activity and greatly lowered electron transfer resistances compared to the unmodified electrodes. Initial proof of concept applications of these electrodes, including the detection of heavy metals by absorptive stripping voltammetry, are also shown.

Arrays of microelectrodes: technologies for environmental investigations.

Within this work it is our intention to provide an overview of the use of arrays or microelectrodes in the characterisation of environmental samples. Electrochemical methods are often a relatively simple and inexpensive alternative to spectroscopic or chromatographic methods for the analysis of a wide range of analytes. Arrays of microelectrodes display a number of advantages over simple planar macroelectrodes and the reasons for this will be detailed within this work. We will also describe some of the most common methods for constructing microarrays. The application of these arrays for analysis of environmental samples such as soil and water for heavy metal contamination has been the major focus of research in this field and comprises much of this review. However other systems will also be detailed such as determination of various anions or other samples such as pesticides.

Label-free impedimetric immunosensors for psoriasin--increased reproducibility and sensitivity using an automated dispensing system.

We describe within this paper the construction of a label-free immunosensor for the protein psoriasin (S100A7), which is associated with a number of clinical conditions such as skin diseases or cancer. Antibodies to psoriasin were immobilised onto screen-printed carbon electrodes that had been pre-modified with the conductive polymer polyaniline. We compared and contrasted a number of different methods of assembly to optimise the construction and properties of the immunosensor. Immunosensors were fabricated using both manual liquid handling (pipette) and an automated liquid dispensing platform, the BioDot AD3200(TM). Two immobilisation methods were also utilised; simple electrostatic binding of the antibody to polyaniline as well as a more complex procedure using a biotin-neutravidin bridge. The optimum results in terms of sensitivity and reproducibility were obtained utilising the automated system and the biotin-avidin assembly procedure. The resultant immunosensors could be interrogated using AC impedance without the need for any labelling and demonstrated quantification of psoriasin from 250 pg ml(-1) to 10 ng ml(-1)-a concentration range suitable for determining physiological levels of psoriasin.

Disposable screen-printed sensors for the electrochemical detection of TNT and DNT.

Due to the heightened level of national security currently prevalent due to the possibility of terrorist incidents, highly portable, miniaturised and sensitive monitoring devices for trace levels of injurious materials, such as explosives are now of the upmost importance. One method that offers a possible route for the development of a detection system for such species is via an electrochemical regime, coupled to the use of disposable sensor technology. Within this study, the use of carbon screen-printed sensors for the detection and analysis of the classical explosive trinitrotoluene (TNT) and the related dinitrotoluene (DNT) is described, with the eventual objective to develop an inexpensive, accurate and sensitive detection system for trace quantities of explosives in field settings. Commercially available screen-printed carbon sensors have been used as the base platform for this investigation and the electrochemistry of both TNT and DNT studied at these surfaces. Two reductive peaks and one oxidative peak were observed for both analytes. The best linear fits and sensitivities were obtained using the reductive peak at -0.72 V vs. Ag/AgCl. A linear range from 1 to 200 µM could be obtained for TNT and DNT in pH 7.0 phosphate buffer with limits of detection as low as 0.4 µM (TNT) and 0.7 µM (DNT). A second system which utilised the addition of the enzyme, nitroreductase, and the coenzyme, NADPH, into the solution matrix prior to electrochemical interrogations with screen-printed carbon electrodes was found to increase the resulting signal magnitude at the oxidation peak at +0.3 V, improving the performance of the sensor at these values.

Detection and imaging the expression of the trans-membrane protein CD44 in RT112 cells by use of enzyme-labeled antibodies and SECM.

The deposition of human RT112 cells in a patterned fashion onto glass substrates and subsequent imaging of the expression of the trans-membrane protein CD44 have been studied using scanning electrochemical microscopy (SECM). Patterns of RT112 cells derived from a transitional cell carcinoma of the bladder could be deposited on amino-modified glass substrates by cytospinning. These were then treated with horseradish peroxidase (HRP) labeled secondary antibodies to the trans-membrane protein CD44. Expression of CD44 protein by the cells directly leads to immobilisation of the labeled antibodies. The presence of the enzyme substrate (hydrogen peroxide) along with a hydroquinone mediator then allowed an enzymatic reaction to proceed, generating benzoquinone. Reduction of benzoquinone gave rise to positive feedback between the substrate and the SECM microelectrode tip. Control samples such as blank slides or slides not treated with HRP-labeled antibody showed negative feedback effects. Patterns of RT112 cells could be assembled and their expression of the target protein imaged whereas control samples showed minimal activity.

Template and catalytic effects of DNA in the construction of polypyrrole/DNA composite macro and microelectrodes.

Electrochemical DNA hybridization-based sensors show great promise as portable and automated analytical devices for routine screening of pathogenic or foreign nucleic acid sequences in biological samples. However, current sensor technologies still exhibit some unresolved issues which hampers their direct application into everyday life. Conducting polymers, such as polypyrrole (PPy), are increasingly being adopted as suitable platforms for DNA probe immobilization and signal transduction. Immobilization of DNA probes during pyrrole electropolymerization is a simple and efficient strategy to build composite electrodes suitable for DNA sensing. However, the effects of the probe state and sequence on PPy growth kinetics have not been studied yet. Here, we show that growth of PPy is drastically affected by the presence of guanine in the DNA probes and whether DNA is present in its single-stranded or double-stranded form. We show that some immobilization protocols may provoke irreversible oxidation of guanine moieties in the probe and that this issue deserves careful investigation as it may interfere with hybridization processes. We have also explored new procedures to build microelectrode arrays bearing immobilized DNA molecules, which are known to show beneficial properties in stirred samples. Overall, we present new techniques and concerns regarding the development of DNA-containing PPy-based composite electrodes, which may be taken into consideration for increasing genosensor reproducibility, response and performance.

A new application of scanning electrochemical microscopy for the label-free interrogation of antibody-antigen interactions: Part 2.

Within this paper we describe the use of scanning electrochemical microscopy (SECM) to fabricate a dotted array of biotinylated polyethyleneimine which was then used to immobilise first neutravidin and then a biotinylated antibody towards a relevant antigen of interest (PSA, NTx, ciprofloxacin). These antigens were selected both for their clinical relevance but also since they display a broad range of molecular weights, to determine whether the size of the antigen used effects the sensitivity of this approach. The SECM was then used to image the binding of both complementary and non-complementary antigens in a label-free assay. Imaging of the arrays before and following exposure to various concentrations of antigen in buffer showed clear evidence for specific binding of the complementary antigens to the antibody functionalised dots. Non-specific binding was also quantified by control experiments with other antigens. This demonstrated non-specific binding across the whole of the substrate, thereby confirming that specific binding does occur between the antibody and antigen of interest at the surface of the dots. The binding of ciprofloxacin was investigated both in simple buffer solution and in a more complex media, bovine milk.

Lactate in human sweat: a critical review of research to the present day.

This review provides a critical overview of the literature published in the area of lactate in human sweat between 1934 and the present. The first section summarizes the relevant pieces of literature, the second evaluates the literature across a range of topics and the third presents potential applications for sweat lactate measurements. Key factors that may affect sweat lactate are discussed in detail in the second section of this review. Both acetylcholine and catecholamine hormonal signals can trigger sweating independently of one another; differences in sweat output and lactate concentrations are compared. The primary triggers for sweating, exercise and heat are also compared, with exercise-induced sweating being further examined with respect to the exercise intensity. This review examines the significant variations in sweat lactate between bodily sites as well the relationship to physiological parameters such as gender, age and physical fitness, along with the effect of climate adaptation on sweat lactate concentrations.

Layer-by-layer immobilization of carbon dots fluorescent nanomaterials on single optical fiber.

We report within this paper the development of a fiber-optic based sensor for Hg(II) ions. Fluorescent carbon nanoparticles were synthesized by laser ablation and functionalized with PEG(200) and N-acetyl-L-cysteine so they can be anionic in nature. This characteristic facilitated their deposition by the layer-by-layer assembly method into thin alternating films along with a cationic polyelectrolyte, poly(ethyleneimine). Such films could be immobilized onto the tip of a glass optical fiber, allowing the construction of an optical fluorescence sensor. When immobilized on the fiber-optic tip, the resultant sensor was capable of selectively detecting sub-micromolar concentrations of Hg(II) with an increased sensitivity compared to carbon dot solutions. The fluorescence of the carbon dots was quenched by up to 44% by Hg(II) ions and interference from other metal ions was minimal.

Recent trends in antibody based sensors.

This review details recent advances in the fields of immunosensors and closely related immunoassays in the past decade, together with a discussion of possible future trends. Immunosensors can be classified by the way in which they transduce the signal produced upon the formation of an antibody antigen complex. Recent advancements to these methods of detection and transduction are discussed in detail, with particular focus on electrochemical, optical, piezoelectric and magnetic based sensors. The varying applications of these sensors are also discussed. Some of the most significant advances include development of immunosensors for the continuous monitoring of analytes, point of care (PoC) devices, with lower unit costs, automation, reusability and ease of use. Immunosensor technology has advanced at a prolific rate since its conception and has grown into a diverse area of ongoing research.

Current trends in explosive detection techniques.

The detection of explosives and explosive-related compounds has become a heightened priority in recent years for homeland security and counter-terrorism applications. There has been a huge increase in research within this area-through both the development of new, innovative detection approaches and the improvement of existing techniques. Developments for miniaturisation, portability, field-ruggedisation and improvements in stand-off distances, selectivity and sensitivity have been necessary to develop and improve techniques. This review provides a consolidation of information relating to recent advances in explosive detection techniques without being limited to one specific research area or explosive type. The focus of this review will be towards advances in the last 5 years, with the reader being referred to earlier reviews where appropriate.

Novel impedimetric immunosensor for the detection and quantitation of Adenovirus using reduced antibody fragments immobilized onto a conducting copolymer surface.

The number of Adenovirus (Ad) infections detected in immunocompromised people has increased due to the number of patients receiving transplants, as well as the HIV pandemic. Ads cause life-threatening diseases specific to the infected organs of immunocompromised hosts, with discontinuation of immunosuppressive agents necessary to prevent morbidity. The methodology in this paper has been employed to develop a novel impedimetric based assay platform to detect and quantify human Ads, which is comparable in performance to current methods, such as ELISA and PCR, but is also less expensive and faster. Novel immunosensors have been fabricated using polyclonal antibodies raised against a human Ad (Ad5) capsid protein, which were selectively cleaved into antibody fragments by 2-mercaptoethylamine. The fragments were immobilized onto a functionalized conducting copolymer matrix comprising polyaniline and 2-aminobenzylamine. Fully fabricated sensors were incubated with two immunologically distinct serotypes of Ad, Ad5 and Ad3, with between 10 and 10(12)virus particles/mL prior to sensor interrogation. Electrochemical impedance spectroscopy was used to measure the charge transfer resistance of the sensors over a range of frequencies from 25 kHz to 0.1 Hz. Our data demonstrate that the immunosensors specifically detect, and differentiate between, closely related human Ad serotypes with a limit of detection of 10(3)virus particles/mL. In addition, atomic force microscopy was applied to study the sensor surface nanostructure. Future work looks to test virus containing clinical samples but this could be a viable and valuable alternative for point-of-care virus detection and quantification.

Construction and interrogation of enzyme microarrays using scanning electrochemical microscopy - optimisation of adsorption and determination of enzymatic activity.

Scanning electrochemical microscopy (SECM) has been used to image and study the catalytic activity of horseradish peroxidase (HRP) immobilised in a patterned fashion onto glass slides. Microarrays of HRP islands could be deposited on amino-modified glass slides using glutaraldehyde crosslinking combined with the SECM being used as a micro-deposition device. The enzymatic activity of the immobilised enzyme on the surface was in the presence of its substrate observed to give rise to substantial positive feedback between the slide and the SECM microelectrode tip. Conversely when either blank slides - or slides coated with HRP which had been subsequently thermally denatured were utilised, these showed negative feedback effects. Various conditions such as enzyme concentration, incubation time and substrate concentration were systematically varied to optimise sensitivity. Regular arrays of HRP could be assembled and when imaged, displayed lower limits of detection of 1.2 × 10(-12) mol ml(-1) of benzoquinone.

A new application of scanning electrochemical microscopy for the label-free interrogation of antibody-antigen interactions.

Within this work we present a 'proof of principle' study for the use of scanning electrochemical microscopy (SECM) to detect and image biomolecular interactions in a label-free assay as a potential alternative to current fluorescence techniques. Screen-printed carbon electrodes were used as the substrate for the deposition of a dotted array, where the dots consist of biotinylated polyethyleneimine. These were then further derivatised, first with neutravidin and then with a biotinylated antibody to the protein neuron specific enolase (NSE). SECM using a ferrocene carboxylic acid mediator showed clear differences between the array and the surrounding unmodified carbon. Imaging of the arrays before and following exposure to various concentrations of the antigen showed clear evidence for specific binding of the NSE antigen to the antibody derivatised dots. Non-specific binding was quantified. Control experiments with other proteins showed only non-specific binding across the whole of the substrate, thereby confirming that specific binding does occur between the antibody and antigen at the surface of the dots. Binding of the antigen was accompanied by a measured increase in current response, which may be explained in terms of protein electrostatic interaction and hydrophobic interactions to the mediator, thereby increasing the localised mediator flux. A calibration curve was obtained between 500 fg mL(-1) to 200 pg mL(-1) NSE which demonstrated a logarithmic relationship between the current change upon binding and antigen concentration without the need for any labelling of the substrate.

Label-free immunochemistry approach to detect and identity antibiotics in milk.

The rise of antibiotic-resistant bacteria is a major concern for the continued health and well-being of the general population. The widespread use of antibiotics within the farming industry is one of the factors that have been linked to the appearance of these resistant strains. Regulation exists to prevent antibiotic foodstuffs such as milk being distributed to consumers; however, for this to be effective, methods must exist for testing the milk to enforce these regulations. Current techniques are often time consuming and expensive, especially when applied to large number of samples. This has led to an interest in milk analyses, which can be carried out outside the laboratory. The development of immunosensors that exploit the exquisite specificity of antibody binding has been studied, both using labeled antibodies such as ELISA assays and, more recently, label-free approaches that directly detect the presence of the antibiotic on binding to a specific antibody. Within this review, we detail recent advances in the detection of antibiotics, especially in milk, using a variety of methods. Various techniques such as electrochemical and surface plasmon resonance methods are described.

Sol-gel based sensor for selective formaldehyde determination.

We report the development of transparent sol-gels with entrapped sensitive and selective reagents for the detection of formaldehyde. The sampling method is based on the adsorption of formaldehyde from the air and reaction with beta-diketones (for example acetylacetone) in a sol-gel matrix to produce a yellow product, lutidine, which was detected directly. The proposed method does not require preparation of samples prior to analysis and allows both screening by visual detection and quantitative measurement by simple spectrophotometry. The detection limit of 0.03 ppmv formaldehyde is reported which is lower than the maximum exposure concentrations recommended by both the World Health Organisation (WHO) and the Occupational Safety and Health Administration (OSHA). This sampling method was found to give good reproducibility, the relative standard deviation at 0.2 and 1 ppmv being 6.3% and 4.6%, respectively. Other carbonyl compounds i.e. acetaldehyde, benzaldehyde, acetone and butanone do not interfere with this analytical approach. Results are provided for the determination of formaldehyde in indoor air.

Sonochemically fabricated microelectrode arrays for use as sensing platforms.

The development, manufacture, modification and subsequent utilisation of sonochemically-formed microelectrode arrays is described for a range of applications. Initial fabrication of the sensing platform utilises ultrasonic ablation of electrochemically insulating polymers deposited upon conductive carbon substrates, forming an array of up to 70,000 microelectrode pores cm(-2). Electrochemical and optical analyses using these arrays, their enhanced signal response and stir-independence area are all discussed. The growth of conducting polymeric "mushroom" protrusion arrays with entrapped biological entities, thereby forming biosensors is detailed. The simplicity and inexpensiveness of this approach, lending itself ideally to mass fabrication coupled with unrivalled sensitivity and stir independence makes commercial viability of this process a reality. Application of microelectrode arrays as functional components within sensors include devices for detection of chlorine, glucose, ethanol and pesticides. Immunosensors based on microelectrode arrays are described within this monograph for antigens associated with prostate cancer and transient ischemic attacks (strokes).

Scanning electrochemical microscopy of genomic DNA microarrays--study of adsorption and subsequent interactions.

The adsorption of genomic DNA and subsequent interactions between adsorbed and solvated DNA have been studied using scanning electrochemical microscopy (SECM). Microarrays of polyethylenimine (PEI) films could be deposited on screen-printed carbon substrates using the SECM. Single stranded herring DNA was electrostatically adsorbed at the surface of the polyethylenimine. The further adsorption of complementary single stranded DNA on the surface was observed to give rise to substantial decreases in interfacial impedance at the surface as measured by increases of tip current of the order of 1-2 nA (6%). Conversely adsorption of DNA from alternate species, i.e. salmon ssDNA on herring ssDNA, yielded much smaller changes in tip current of 0.2 nA. The significance of this work is that the approach opens up the possibility for direct label-free electrochemical interrogation of DNA microarrays as an alternative to other existing optical techniques.