Editorial, Seminars in Cell & Developmental Biology

It is a pleasure to introduce this special edition of Cell and Development Biology dedicated to the field and application of Biosensors. This edition comprises seven reviews covering the most active research areas where we believe some of the most prominent advances in the field are likely to emerge in the near to medium term. In line with scope of this journal, some emphasis is given towards techniques applicable to Cell Biology

Carrier systems and biosensors for biomedical applications.

This chapter addresses both carrier systems and biosensors which are often applied directly to tissues, either as skin patches, implanted or ingested by a variety of routes. It follows that there is a common theme between these applications and many of those discussed elsewhere within this book. Any device, scaffold or implant within the body must usually display extreme biocompatibility if it is not to cause harm to the patient. The techniques of tailoring surfaces to ensure no adverse reactions are a common theme running throughout this work on tissue engineering

Sonochemically fabricated microelectrode arrays for biosensors. Part II. Modification with a polysiloxane coating

A polymer modified sonochemically fabricated glucose oxidase microelectrode array with microelectrode population densities of up to 2.5 x 105 microelectrodes cm-2 is reported. These microelectrode sensors were formed by first depositing an insulating film on commercial screen printed electrodes which was subsequently sonicated to form cavities of regular sizes in the film. Electropolymerisation of aniline at the microelectrode cavities formed polyaniline protrusions containing entrapped glucose oxidase. Chemical deposition of polysiloxane from dichlorodimethysilane was used to deposit a thin protective and diffusion mass transport controlling coating over the electrodes. The physical and electrochemical properties of these films were studied. The performance of the final glucose oxidase based microelectrode sensor array is reported

Flexible Ultrathin PolyDVB/EVB Composite Membranes for the Optimization of a Whole Blood Glucose Sensor.

An ultrathin composite membrane has been developed as the outer covering barrier in a model amperometric glucose oxidase enzyme electrode. The membrane was formed by cathodic electropolymerization of divinylbenzene/ethylvinylbenzene at the surface of a gold coated polyester support membrane. Permeability coefficients were determined for O2 and glucose across membranes with a range of polymer thicknesses. Anionic interferents (such as ascorbate), were screened from the working electrode via a charge exclusion mechanism. The enzyme electrode showed an initial 10% signal drift when first exposed to whole human blood over a period of 2 hours, after which responses remained essentially stable. Whole blood patient glucose determinations yielded a correlation coefficient of r2=0.99 compared to standard hospital analyses

Electrochemical detection of TNT at cobalt phthalocyanine mediated screen-printed electrodes and application to detection of airborne vapours

We describe the use of cobalt phthalocyanine as a mediator to improve the sensitivity for the electrochemical detection of TNT. Commercial screen-printed electrodes containing cobalt phthalocyanine were employed for determination of TNT. Improved sensitivities compared to screen-printed carbon electrodes without phthalocyanine were observed, current response for cyclic voltammetric measurements at modified electrodes being at least double that of unmodified electrodes. A synergistic effect between oxygen and TNT reduction was also observed. Correlation between TNT concentrations and sensor output was observed between 0–200 µM TNT. Initial proof-of-concept experiments combining electrochemical determinations, with the use of an air-sampling cyclone, are also reported

Scanning electrochemical microscopy for the characterisation of surfaces modified with biological molecules

This thesis describes a novel fabrication procedure for microelectrodes to be used with the scanning electrochemical microscope (SECM), the characterisation of a variety of novel impedance based immunosensors, and the characterisation of a novel oligonucleotide biosensor. The thesis firstly describes the development of a protocol for the fabrication of reproducible microelectrodes characterised to identify suitability in use with the SECM. The thesis then describes the use of SECM in feedback mode to interrogate a variety of antibody-polyelectrolyte films determining whether the changes observed by impedance were detectable by SECM. A screen printed carbon ink surface was patterned with an array of biotinylated polyethyleneimine (PEI) which was exposed to Neutravidin and then the biotinylated antibody of interest. Using ferrocenecarboxylic acid as the redox couple, the array was interrogated by SECM, scanning before and following exposure to a series of concentrations of the complementary antigen and a non-complementary antigen. Upon the exposure of the PEI/Neutravidin/biotinylated antibody array to the antigen, the feedback current over the functionalised region was observed to change. The change observed increased as the concentration of the antigen exposed to the array was increased showing linear correlation. On exposure of the array to a non-complementary antigen, only a small change in the feedback current was observed. NSE, PSA, Ciprofloxacin and NTx were all investigated with limits of detection of 0.5 pg ml-1, 1 pg ml-1, 0.1 ng ml-1 and 1 nM respectively. Finally using a similar method as employed above, SECM was utilised in the detection of binding events of short oligonucleotides. Once again scans were conducted before and after exposure to both complementary and non-complementary oligonucleotide sequences and by subtraction absolute changes in feedback current were determined. On exposure to the complementary oligonucleotide sequence a change in feedback was observed when the array was exposed to the non-complementary oligonucleotide sequence, as with the antibody/antigen array, only a small change in the feedback current was observed.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Studies towards the exploitation of sonochemically formed microelectrode arrays for the development of electrochemical sensors

Microelectrodes offer a number of advantages for exploitation as electrochemical sensors such as imparting stir-independence to sensor responses and allowing lower limits of detection to be minimised. Microelectrode arrays offer an attractive route for increasing the current responses of microelectrodes, whilst still retaining their advantageous properties. Despite this, no commercial sensors, to date, have successfully employed microelectrode arrays, largely due to conventional fabrication routes proving too costly to be economically viable for the production of disposable sensing devices. Previous work carried out by this research group has described a novel and patented procedure for the fabrication of microelectrode arrays via the sonochemical ablation of insulating polymer films electrochemically deposited upon conductive surfaces. This format lends itself to mass fabrication due to the simplicity and inexpensiveness of the approach. This thesis describes work focussed towards the optimisation of each of the individual components involved in the formation of sonochemically fabricated microelectrode arrays. In particular, factors and techniques that may facilitate the commercial exploitation and mass fabrication of such arrays as generic sensing templates are described. Screen printed carbon has been investigated for its suitability as a host electrode. The comparative use of a number of possible activation methods to increase amperometric current responses at such electrodes is also described. Homogeneous poly(o-phenylenediamine) films of -40 nm thickness formed at the surfaces of screen printed carbon electrodes via the anodic electropolymerisation of o-phenylenediamine are shown to serve as effective diffusional barriers, thus insulating the underlying carbon electrodes. Microelectrode arrays formed by the sonochemical ablation of such films to expose microscopic areas of the underlying conductive substrates are seen to possess electrode element populations of -7.3 x 104 cm 2. Over 400 such sensors are shown to be able to be fabricated simultaneously with reproducibility of responses <4% relative standard deviation. Amperometric and cyclic voltammetric characterisations of the thus produced microelectrode arrays performed in model redox systems are shown to agree with accepted theoretical microelectrode behaviour, demonstrating sigmoidal shaped voltammograms, fluctuations in steady-state current responses of <10% with convection, scan rate independence and fast attainment (<20 seconds) of steady-state responses. Arrays of this type are also demonstrated to be suitable for exploitation within aqueous chlorine sensing devices, offering detection limits of <0.005 mg/l free chlorine, representing an order of magnitude lower than those obtainable via contemporary optical wet chemistry based approaches. In order to demonstrate further the applicability of this approach to the mass fabrication of disposable devices, methods for the deposition of a chemical modifying layer are also investigated, to avoid the need for additional reagents.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Development of impedimetric immunosensors for the detection of a range of antigens of biological or biomedical significance

This PhD project was funded by the European Union Framework 6 ELISHA programme as part of the ELISHA programme. The aim of the research was focused towards the development of point-of-care, simple, cost-efficient and reliable impedimetric immunosensors for the rapid detection of important antigens such as ciprofloxacin and digoxin. Through the cooperation of the 9 involved partners a number of protocols have been developed for sensor fabrication and sample testing that allow both rapid and reliable detection of a range of antigens. This work describes in depth, the use of polymers and cyclic voltammetry for electrode surface modification, the use of the avidin-biotin system for antibody immobilisation and finally the use of Electrochemical Impedance Spectroscopy for antigen detection. We report here the successful development of electrochemical impedimetric carbon based immunosensors for the detection of free-form and chelated ciprofloxacin (both in laboratory buffer and milk), digoxin and green fluorescent protein. It was observed during this work that unavailability of sufficient quantities of the monoclonal antibodies could lead to early sensor saturation and hence a less extended antigen detection range, while very low quantities of immobilised antibodies could also give rise to erroneous results. The immunosensors towards ciprofloxacin detected the respective antigen when this was present between 1 ng ml-1 and 10 μg ml-1 in PBS buffer and 0.1 ng ml-1 to 10 μg ml-1 when the antigen was added to milk samples. The developed digoxin immunosensors could detect digoxin between the concentrations of 0.1 ng ml-1 and 10 μg ml-1. Lower detection limits where observed for the sensors targeting GFP which could detect their respective antigen at concentrations as low as 100 pg ml-1. The tested concentration range of the latter sensors was extended up to 100 ng ml-1.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Towards a commerical microelectrode array based sensor for improved chlorine detection

The commercial development of a disposable aqueous chlorine sensor based on a novel microelectrode array fabrication process is described. Non-conducting poly(o-phenylenediamine) films are firstly used to passivate conductive surfaces. Ultrasonic ablation of passivated electrode assemblies then results in the formation of a plurality of wells to expose the underlying conductive substrate, thereby forming a microelectrode array. Microelectrode arrays produced in this manner can be exploited within many electrochemical sensing applications; however, portable aqueous chlorine detection has been selected by Microarray Limited (the industrial sponsors of this project) as a primary vehicle for launching its generic production technology. The scale of microelectrode array production has been extended from that of individual gold sputtercoated glass slide electrodes - to the simultaneous production of hundreds of low-cost screen printed carbon-ink based sensors. A focus has been directed at all stages towards permitting the cost-effective large-scale mass production of sensors with a view to challenging existing portable aqueous chlorine measurement technologies both in terms of performance and unit cost. Based on volume batches of 250,000, it has been calculated that Microarray Limited sensors can be manufactured for a unit cost of approximately 2.5 pence, sufficiently low to provide scope for a competitive yet profitable sale price. The Microarray Limited aqueous chlorine detection system has improved the limit of detection from 0.01 ppm to 0.005 ppm total chlorine without sacrificing accuracy. Furthermore, this novel approach to aqueous chlorine detection offers numerous key benefits to the customer including reduced testing time, a more straightforward operation and the elimination of harmful reagents. Product development has been described from an initial concept through to a pre-production phase. The development of an innovative generic sensor packaging technology is also described.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Labeless and reversible immunosensor assay based upon an electrochemical current-transient protocol

A novel labeless and reversible immunoassay based upon an electrochemical current-transient protocol is reported which offers many advantages in comparison to classical immuno-biochemical analyses in terms of simplicity, speed of response, reusability and possibility of multiple determinations. Conducting polypyrrole films containing antibodies against 1) Bovine Serum Albumin (BSA) and 2) Digoxin were deposited on the surface of platinum electrodes to produce conductive affinity matrices having clearly defined binding characteristics. The deposition process has been investigated using 125I labelled anti-digoxin to determine optimal fabrication protocols. Antibody integrity and activity, together with non-specific binding of antigen on the conducting matrix have also been investigated using tritiated digoxin to probe polypyrrole/anti-digoxin films. Amperometric responses to digoxin were recorded in flow conditions using these films, but the technique was limited in use mainly due to baseline instability. Anti-BSA - polypyrrole matrices were investigated in more detail in both flow and quiescent conditions. No observable response was found in flow conditions, however under quiescent conditions (in non-stirred batch cell), anti-BSA – polypyrrole films have been demonstrated to function as novel quantitative chronoamperometric immuno-biosensors when interrogated using a pulsed potential waveform. The behaviour of the electrodes showed that the antibody/antigen binding and/or interaction process underlying the response observed was reversible in nature, indicating that the electrodes could be used for multiple sensing protocols. Calibration profiles for BSA demonstrated linearity for a concentration range of 0-50 ppm but tended towards a plateau at higher concentrations. Factors relating to replicate sensor production, sample measurement and reproducibility are discuss