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Electrochemical sensing is ubiquitous in a number of fields ranging from biosensing, to environmental monitoring through to food safety and battery or corrosion characterisation. Whereas conventional potentiostats are ideal to develop assays in laboratory settings, they are in general, not well-suited for field work due to their size and power requirements. To address this need, a number of portable battery-operated potentiostats have been proposed over the years. However, most open source solutions do not take full advantage of integrated circuit (IC) potentiostats, a rapidly evolving field. This is partly due to the constraining requirements inherent to the development of dedicated interfaces, such as apps, to address and control a set of common electrochemical sensing parameters. Here we propose the PocketEC, a universal app that has all the functionalities to interface with potentiostat ICs through a user defined property file. The versatility of PocketEC, developed with an assay developer mindset, was demonstrated by interfacing it, via Bluetooth, to the ADuCM355 evaluation board, the open-source DStat potentiostat and the Voyager board, a custom-built, small footprint potentiostat based around the LMP91000 chip. The Voyager board is presented here for the first time. Data obtained using a standard redox probe, Ferrocene Carboxylic Acid (FCA) and a silver ion assay using anodic stripping multi-step amperometry were in good agreement with analogous measurements using a bench top potentiostat. Combined with its Voyager board companion, the PocketEC app can be used directly for a number of wearable or portable electrochemical sensing applications. Importantly, the versatility of the app makes it a candidate of choice for the development of future portable potentiostats. Finally, the app is available to download on the Google Play store and the source codes and design files for the PocketEC app and the Voyager board are shared via Creative Commons license (CC BY-NC 3.0) to promote the development of novel portable or wearable applications based on electrochemical sensing.
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Antimicrobial impregnated wound dressings are a critical tool for the management, prevention, and control of surgical site infections (SSIs) and infected chronic wounds. However, the sustained therapeutic antimicrobial activity of the dressing when employed for extended periods cannot be readily determined in vivo. Consequently, dressings are changed frequently to ensure that their antimicrobial activity is maintained. Whilst frequent dressing changes allow the wound to be assessed, this is time-consuming and can cause disruption to the wound bed impairing the healing process. Furthermore, this increases medical costs for the patient and hospitals. This paper introduces a novel concept to monitor the therapeutic levels of an antimicrobial component within a wound dressing ensuring the wound dressing remains "fit for purpose" and avoiding indiscriminate use of antiseptics. This could help to inform clinicians whether the antimicrobial is still being delivered at therapeutic levels and as such when to change the dressing ensuring timely positive clinical outcomes. Silver has been used historically as an antimicrobial agent and is ubiquitous in current generations of antimicrobial wound dressings. However, its activity is complex due to the poor solubility of silver ions in the presence of chloride and the effect of complexation by other components in the dressing and wound ecosystem, not least by serum proteins. In this paper, we detail an electrochemical silver sensor (5D patent protected - WO2023275553A1), constructed using a platinum (Pt) nanoband array electrode, and characterise its response to silver ions. This is determined in the presence of bovine serum albumin (BSA) and simulated wound fluid (SWF) containing chloride and rationalised using atomic analysis of the composition of the SWF. The sensor response in SWF is compared with the antimicrobial activity of silver against Pseudomonas aeruginosa in the planktonic and biofilm state, as a function of the amount of silver nitrate added. At low concentrations, silver in SWF has good solubility but reduced antimicrobial effect due to binding of silver by BSA as shown by the sensor response. At intermediate concentrations, above 10ppm, the silver was efficacious on both planktonic microorganisms and biofilm impregnated with microorganisms and readily detected with the sensor. At high concentrations, silver precipitates and both the silver in solution and the sensor response plateaus. The data demonstrates how the sensor correlates with the antimicrobial activity of the silver in vitro and how this could be used to actively monitor antimicrobials in vivo.
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The performance of two electrode architectures with broadly similar overall active electrode areas are examined. The first is an electrode comprising a single contiguous area (a disc) and the second is an electrode in which the cumulative electrode area is dispersed over a wide area as a 50 nm thickness platinum nanoband. A direct comparison of the electrochemical performance of these two electrodes has been made. The relatively simple nanoband electrode architecture is shown to have benefits, including two orders of magnitude greater mass transport limited currents, the ability to measure faster electrode kinetics (by a similar factor), a three orders of magnitude lowering of the Limit of Detection and a significantly reduced susceptibility to hydrodynamic perturbations. The consequences and implications of these performance characteristics on the uses of such a nanoband electrode have been considered.
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Background: Assessment of paracetamol overdose in children and teenagers in the emergency department (ED) requires blood, taken 4 hours post ingestion. A commercial partner developed transdermal paracetamol measuring technology. This work aims to understand the acceptability of such a device, and potential market size. Methods: A questionnaire study was undertaken with children and parents attending Alder Hey Children's Hospital, and healthcare professionals (HCP) involved in their care. A retrospective audit of paracetamol ingestion presenting to a paediatric ED was undertaken. Results: One hundred forty-three questionnaires were distributed, and 139 returned (response rate 97.2%), comprising 55 children, 52 parents and 32 HCP (recruited between August-October 2019). Overall device acceptability, assessed by favourability of appearance and willingness to wear was high, at 60.0% and 81.5% respectively. Concerns raised included bulky size and weight, and concern regarding the duration younger children would tolerate wearing the device. All groups, including children, ranked accuracy of results as the most important device feature and device comfort the least important. Parents prioritised avoidance of blood tests more than children. One hundred twenty-seven children presented to ED with paracetamol ingestion (September 2017-August 2018), with 57 (44.9%) categorised as accidental overdose. Overall, 106 (83.4%) required paracetamol concentration measuring, and 25 (19.7%) of these required treatment with N-acetylcysteine. Extrapolating nationally, over 7000 children will present with accidental overdose per annum in the UK. Conclusion: Acceptability of a non-invasive paracetamol sensor was high in all groups, provided accuracy could be assured.
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Dual polarisation interferometry (DPI) has been used to characterise the formation of hybrid bilayer membranes (HBM) on a silicon-oxynitride surface. This technique allows the simultaneous determination of multiple physical properties of an HBM, as the HBM is being formed in a single experiment: mass, thickness in the z-direction (normal to the surface), tilt angle of the first layer and refractive index. Decanoic acid was covalently attached to an amine modified silicon-oxynitride sensor chip surface via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride condensation reaction. The decanoic acid layer was 0.92+/-0.12 nm thick, indicating a tilt angle of 57 degrees from surface normal, and possessed a mass of 1.05+/-0.10 ng mm(-2) and a refractive index (RI) of 1.450+/-0.020. Phospholipid vesicles made from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) or 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) were injected over the fatty acid surface to form an HBM. The DPPC HBM was 4.32+/-0.68 nm thick, with a total mass of 3.18+/-0.60 ng mm(-2) and a RI of 1.404+/-0.007. The DMPC HBM was 2.12+/-0.34 nm thick, with a total mass of 2.25+/-0.51 ng mm(-2), and a RI of 1.435+/-0.007. DPI thus provides an insight into HBM formation and differences between the structural organisation of HBMs of different composition.
Assuntos
Materiais Biomiméticos/química , Bicamadas Lipídicas/química , Lipossomos/química , Teste de Materiais/métodos , Fluidez de Membrana , Microscopia de Interferência/métodos , Microscopia de Polarização/métodos , Materiais Biomiméticos/análise , Bicamadas Lipídicas/análise , Lipossomos/análise , Membranas Artificiais , Microscopia de Interferência/instrumentação , Microscopia de Polarização/instrumentaçãoRESUMO
We describe a novel glucose biosensor based on a nanoband array electrode design, manufactured using standard semiconductor processing techniques, and bio-modified with glucose oxidase immobilized at the nanoband electrode surface. The nanoband array architecture allows for efficient diffusion of glucose and oxygen to the electrode, resulting in a thousand-fold improvement in sensitivity and wide linear range compared to a conventional electrode. The electrode constitutes a robust and manufacturable sensing platform.
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Técnicas Biossensoriais/métodos , Glucose Oxidase/metabolismo , Glucose/análise , Nanotecnologia/métodos , Técnicas Biossensoriais/instrumentação , Eletroquímica , Eletrodos , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Glucose/química , Glucose Oxidase/química , Limite de Detecção , Nanotecnologia/instrumentação , Platina/química , Propriedades de SuperfícieRESUMO
Apolipoprotein E (Apo E) is an important genetic risk factor for multiple neurological, vascular and cardiovascular diseases. Previously, we reported Apo E isoprotein-specific modulation of tissue plasminogen activator (tPA) using an in vitro blood-clotting assay. Here, we studied the conformational changes of Apo E2, E3 and E4 in the presence of tPA and vice versa using circular dichroism (CD) and dual polarization interferometry (DPI). We report isoprotein and state-specific intermolecular interactions between the Apo E isoforms and tPA. Apo E2 interaction with immobilized tPA leads to significant conformational changes which are not observed with Apo E3 or E4. Additionally, tPA induces changes in helicity of lipidated Apo E2 whereas no detectable changes were observed in Apo E3 or E4. The Tukey's test for interaction indicated a significant (P < 0.001) interaction between tPA and Apo E2 in the lipidated environment. These results may be important regarding the mechanism by which Apo E has isoprotein-specific effects on many biological processes and diseases involving blood clotting, proteolysis and perfusion.
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Apolipoproteínas E/metabolismo , Ativador de Plasminogênio Tecidual/metabolismo , Humanos , Ligação ProteicaRESUMO
A new optical biosensor is described based on a dual waveguide interferometric technique. By addressing the waveguide structure with alternate polarisations the optogeometrical properties (density and thickness) of adsorbed protein layers at the sensor (solid)-liquid interface have been determined. Differences in the waveguide mode dispersion between the transverse electric (TE) and transverse magnetic (TM) modes allow unique solutions for adlayer thickness and refractive index to be determined at all stages during the formation process. The technique has been verified using standard protein systems and by comparing the data with published work using X-ray crystallography and neutron reflection techniques. The data obtained was found to be in excellent agreement with previously reported X-ray experiments given that typical film thicknesses for streptavidin layers were in the range 5.5-6.5 nm compared with the short axis crystal structure of between 4.8 and 5.6 nm. The precision of the measurements taken was of the order of 40 pm with respect to adsorbed adlayer thicknesses. This biosensor approach provides measurements of both thickness and density of adlayers to a high precision, simultaneously and in real time enabling detail of the structure and function of proteins to be elucidated. From such data it is possible to obtain information on the orientation, distortion and efficiency of immobilisation procedures as well as the interaction event of interest. The technique is expected to find utility with those interested in protein structure and function. This is an area of growing importance within the life sciences as the demand for quantitative analytical techniques increases with the growth in "proteomics".
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Técnicas Biossensoriais/instrumentação , Análise de Falha de Equipamento , Interferometria/instrumentação , Óptica e Fotônica/instrumentação , Proteínas/análise , Proteínas/química , Anticorpos/análise , Anticorpos/química , Técnicas Biossensoriais/métodos , Biotina/análise , Biotina/química , Desenho de Equipamento , Humanos , Interferometria/métodos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Estreptavidina/análise , Estreptavidina/químicaRESUMO
Micron resolution photolithography has been employed to make microsquare nanoband edge electrode (MNEE) arrays with reproducible and systematic control of the crucial dimensional parameters, including array element size and spacing and nanoelectrode thickness. The response of these arrays, which can be reproducibly fabricated on a commercial scale, is first established. The resulting characteristics (including high signal and signal-to-noise, low limit of detection, insensitivity to external convection and fast, steady-state, reproducible and quantitative response) make such nanoband electrode arrays of real interest as enhanced electroanalytical devices. In particular, the nanoelectrode response is presented and analysed as a function of nanometre scale electrode dimension, to assess the impact and relative contributions of previously postulated nanodimensional effects on the resulting response. This work suggests a significant contribution of migration at the band edges to mass transfer, which affects the resulting electroanalytical response even at ionic strengths as large as 0.7 mol dm(-3) and for electrodes as wide as 50 nm. For 5 nm nanobands, additional nanoeffects, which are thought to arise from the fact that the size of the redox species is comparable to the band width, are also observed to attenuate the observed current. The fundamental insight this gives into electrode performance is discussed along with the consequent impact on using such electrodes of nanometre dimension.
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A novel technique for the production of nanoscale electrode arrays that uses standard microfabrication processes and micron-scale photolithography is reported here in detail. These microsquare nanoband edge electrode (MNEE) arrays have been fabricated with highly reproducible control of the key array dimensions, including the size and pitch of the individual elements and, most importantly, the width of the nanoband electrodes. The definition of lateral features to nanoscale dimensions typically requires expensive patterning techniques that are complex and low-throughput. However, the fabrication methodology used here relies on the fact that vertical dimensions (i.e. layer thicknesses) have long been manufacturable at the nanoscale using thin film deposition techniques that are well established in mainstream microelectronics. The authors report for the first time two aspects that highlight the particular suitability of these MNEE array systems for probe monolayer biosensing. The first is simulation, which shows the enhanced sensitivity to the redox reaction of the solution redox couple. The second is the enhancement of probe film functionalisation observed for the probe film model molecule, 6-mercapto-1-hexanol compared with microsquare electrodes. Such surface modification for specific probe layer biosensing and detection is of significance for a wide range of biomedical and other sensing and analytical applications.
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Técnicas Biossensoriais/instrumentação , Nanotecnologia/métodos , Algoritmos , Simulação por Computador , Eletroquímica/métodos , Eletrodos , Desenho de Equipamento , Análise de Elementos Finitos , Nanoestruturas , Oxirredução , Propriedades de SuperfícieRESUMO
Dual polarization interferometry (DPI) is an analytical technique that allows the simultaneous determination of thickness, density, and mass of a biological layer on a sensing waveguide surface in real time. The technique was applied to the analysis of carbohydrate-protein interactions. The selected system involved a 12-kDa recombinant fragment of collagen V (HepV) and heparin, a complex polysaccharide. Here we report on the analysis of thickness, density, and mass of surface structures obtained during the binding of HepV to heparin, which is a useful model compound for the sulfated, protein-binding regions of heparan sulfate. This system, which was initially studied for its biological relevance, displayed anomalous behavior in kinetic studies using surface plasmon resonance (SPR) assays that has been attributed to putative conformational changes. It was this putative conformational change that prompted us to investigate the binding using an alternative analytical approach. While using DPI to monitor binding events, a streptavidin layer (surface coverage 2.105 ng mm(-2)) was bound to the sensor surface (92% coverage), which captured 0.105 ng mm(-2) of biotinylated heparin (a stoichiometric ratio of 1:6 heparin-streptavidin). The heparin inserted into the streptavidin layer but was still found to be capable of binding 0.154 ng mm(-2) of HepV, which was also observed to insert into the streptavidin layer. This allowed the reliable calculation of the stoichiometric ratio for the HepV-heparin complex ( approximately 1.7:1.0), which has proved to be difficult to evaluate by SPR assays. Furthermore, real-time analysis of the heparin-HepV interaction by DPI suggested that there was some surface loss (probably of streptavidin) while the binding was occurring rather than the putative conformational change that has been suggested on the basis of kinetic data alone. This gives further insight into the binding mechanism of HepV to heparin.
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Colágeno Tipo V/química , Heparina/química , Sítios de Ligação , Biotinilação , Colágeno Tipo V/análise , Heparina/análise , Interferometria/métodos , Cinética , Ligação Proteica , Proteínas Recombinantes/análise , Proteínas Recombinantes/química , Sensibilidade e Especificidade , Estreptavidina/química , Ressonância de Plasmônio de Superfície/métodos , Propriedades de Superfície , Fatores de TempoRESUMO
The study of solution-phase interactions between small molecules and immobilized proteins is of intense interest, especially to the pharmaceutical industry. An optical sensing technique, dual polarization interferometry, has been employed for the detailed study of a model protein system, namely, d-biotin interactions with streptavidin immobilized on a solid surface. Changes in thickness and density of an immobilized streptavidin layer as a result of the binding of d-biotin have been directly measured in solution and in real time. The results obtained from this approach are in excellent agreement with X-ray crystallographic data for the structural changes expected in the streptavidin-D-biotin system. The mass changes measured on binding d-biotin also agree closely with anticipated binding capacity values. Determination of the density changes occurring in the protein adlayer provides a means for differentiation between specific and nonspecific interactions.
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Biotina/metabolismo , Técnicas de Química Analítica/métodos , Estreptavidina/química , Interferometria/métodos , Luz , Ligação Proteica , Estrutura Terciária de Proteína , Estreptavidina/metabolismoRESUMO
Lysozyme adsorption at the silica/water interface has been studied using a new analytical technique called dual polarization interferometry. This laboratory-based technique allows the build up or removal of molecular layers adsorbing or reacting on a lightly doped silicon dioxide (silica) surface to be measured in terms of thickness and refractive index changes with time. Lysozyme adsorption was studied at a range of concentrations from 0.03 to 4.0 g dm(-3) and at both pH 4 and pH 7. Adsorbed layers ranging from 14 to 43 +/- 1 A in thickness and 0.21 to 2.36 +/- 0.05 mg m(-2) in mass coverage were observed at pH 4 with increasing lysozyme concentration, indicating a strong deformation of the monolayer over the low concentration range and the formation of an almost complete sideways-on bilayer toward the high concentration of 4 g dm(-3). At pH 7, the thickness of adsorbed layers varied from 16 to 54 +/- 1 A with significantly higher surface coverage (0.74 to 3.29 +/- 0.05 mg m(-2)), again indicating structural deformation during the initial monolayer formation, followed by a gradual transition to bilayer adsorption over the high concentration end. The pH recycling performed at a fixed lysozyme concentration of 1.0 g dm(-3) indicated a broadly reversible adsorption regardless of whether the pH was cycled from pH 7 to pH 4 and back again or vice versa. These observations are in good agreement with earlier studies undertaken using neutron reflection although the fine details of molecular orientations in the layers differ subtly.