A fully integrated microfluidic device for point of care monitoring of antithrombotics

© 2016 The Royal Society of Chemistry. The simplicity and efficiency of point of care diagnostics have revolutionised patient care. Current methods for measuring hypercoagulability often require trained technicians, large blood volumes, and result in long turnaround times. Standard testing for hypercoagulable disorders is performed in the central laboratory using automated coagulation analysers. However the trend is moving towards the development and implementation of point of care testing, as a result of the ever increasing number of patients on antithrombotic therapy. We present a novel microfluidic device and assay for monitoring the effect of two anticoagulants, unfractionated heparin (UFH) and low molecular weight heparin (LMWH). The assay is based on the anti-Xa assay principle but uses fluorescence detection. Our device is a disposable laminate microfluidic strip, fabricated from the cyclic polyolefin (COP), Zeonor®, which is extremely suitable for application to fluorescent device platforms. We present data on the execution of the anti-Xa assay in this microfluidic format, demonstrating that the assay can be used to measure both UFH and LMWH in human plasma samples from 0 to 1 U mL-1, with a rapid result obtained within 30-60 seconds

Development and characterization of nickel-NTA-polyaniline modified electrodes

The engineered addition of hexa-histidine sequences to biomolecules such as antibody fragments has been found to be an excellent means of purifying these materials. This tagging methodology has also been extended to its use as a tool for immobilization and orientation of antibodies on transducer surfaces. Polyvinyl sulfonate-doped polyanilne (PANI/ PVS) can be used as a mediator in amperometric biosensors. This short communication looks at the effect of nickel chelate materials and nickel chelation on this conducting polymer and evaluates it as a potential surface for the immobilization of his-tagged biomolecules. N-nitrilotriacetic acid (NTA) was doped into the electropolymerized PANI/PVS at a screen-printed carbon paste electrode. The resulting NTA-PANI/PVS film was shown to have comparable electrochemical properties of polymer without the chelating agent. When Ni 2þ was applied to the electrode, the incorporated NTA was found to efficiently chelate the metal ions at the electrode surface

Electrochemistry and application of a novel monosubstituted squarate electron-transfer mediator in a glucose oxidase-doped poly(phenol) sensor

Electrosynthetic poly(phenol) nanofilms were deposited in situ on platinum electrodes in the presence and absence of glucose oxidase. The synthesis charges and currents of the nonconducting polymer films were recorded at various applied potentials for films grown from 25–100 mM phenol concentrations. Film parameters such as the standard rate constant for film deposition, film thickness, and surface concentration of the poly(phenol) films were evaluated from the cyclic and step voltammograms of the polymerization process. A novel electron-transfer mediator consisting of monosubstituted 4-hydroxycyclobut-3-ene-1,2-dione (squarate) was used as a mediator for Pt/poly(phenol) nano-film/GOx amperometric glucose biosensors. Amperometric responses for 3-diphenylamino-4-hydroxycyclobut-3-ene-1,2- dione (diphenylaminosquarate: E°′ = of +328 mV/Ag-AgCl at pH 7.0)-mediated systems were measured by both steady-state amperometric and cyclic voltammetry. The sensor sensitivity was calculated to be 558 nA cm –2 (µM) –1

Optimisation and characterisation of biosensors based on polyaniline

With lower limits of detection and increased stability constantly being demanded of biosensor devices, characterisation of the constituent layers that make up the sensor has become unavoidable, since this is inextricably linked with its performance. This work describe the optimisation and characterisation of two aspects of sensor performance: a conductive polymer layer (polyaniline) and the immobilised protein layer. The influence of the thickness of polyaniline films deposited electrochemically onto screen-printed electrode surfaces is described in this work in terms of its influence on a variety of amperometric sensor performance characteristics: time to reach steady state, charging current, catalytic current, background current and signal/background ratios. The influence of polymer film thickness on the conductivity and morphology of finished films is also presented. An electrostatic method of protein immobilisation is used in this work and scanning electron microscopy in conjunction with gold-labelled antibodies and back-scattered electron detection has enabled the direct visualisation of individual groups of proteins on the sensor surface. Such information can provide an insight into the performance of sensors under influence of increasing protein concentrations

Application of nanoparticulate conducting polyaniline in nanofilm biosensor technology

This biosensor uses a novel aqueous-based nanoparticulate polyaniline (PANI), synthesised using dodecylbenzenesulphonic acid (DBSA) and aniline as starting material. These polymer nanoparticles have been electrodeposited on to the surface of carbon electrodes resulting in conductive nano-films, which were examined by electrochemistry, scanning electron microscopy (SEM), atomic force microscopy (AFM), profilometry and spectroelectrochemistry. Biomolecules were then electrostatically adsorbed onto this surface and physical techniques have shown that the nanofilm possesses properties which allow for uniform adsorption of protein to take place. This effective biosensor format has been characterised using a horseradish peroxidase (HRP) and H2O2 format. This sensor exhibits higher signal/noise (S/N) ratios and quicker response times than previous PANI biosensor formats developed by our group, due to its nanofilm characteristic

Disposable sensors in diagnostics, food and environmental monitoring

Disposable sensors are low‐cost and easy‐to‐use sensing devices intended for short‐term or rapid single‐point measurements. The growing demand for fast, accessible, and reliable information in a vastly connected world makes disposable sensors increasingly important. The areas of application for such devices are numerous, ranging from pharmaceutical, agricultural, environmental, forensic, and food sciences to wearables and clinical diagnostics, especially in resource‐limited settings. The capabilities of disposable sensors can extend beyond measuring traditional physical quantities (for example, temperature or pressure); they can provide critical chemical and biological information (chemo‐ and biosensors) that can be digitized and made available to users and centralized/decentralized facilities for data storage, remotely. These features could pave the way for new classes of low‐cost systems for health, food, and environmental monitoring that can democratize sensing across the globe. Here, a brief insight into the materials and basics of sensors (methods of transduction, molecular recognition, and amplification) is provided followed by a comprehensive and critical overview of the disposable sensors currently used for medical diagnostics, food, and environmental analysis. Finally, views on how the field of disposable sensing devices will continue its evolution are discussed, including the future trends, challenges, and opportunities

Measurement of the viscoelastic properties of blood plasma clot formation in response to tissue factor concentration-dependent activation

© 2016, The Author(s). The coagulation of blood plasma in response to activation with a range of tissue factor (TF) concentrations was studied with a quartz crystal microbalance (QCM), where frequency and half width at half maximum (bandwidth) values measured from the conductance spectrum near resonant frequency were used. Continuous measurement of bandwidth along with the frequency allows for an understanding of the dissipative nature of the forming viscoelastic clot, thus providing information on the complex kinetics of the viscoelastic changes occurring during the clot formation process. Using a mathematical model, these changes in frequency and bandwidth have been used to derive novel QCM parameters of effective elasticity, effective mass density and rigidity factor of the viscoelastic layer. The responses of QCM were compared with those from thromboelastography (TEG) under identical conditions. It was demonstrated that the nature of the clot formed, as determined from the QCM parameters, was highly dependent on the rate of clot formation resulting from the TF concentration used for activation. These parameters could also be related to physical clot characteristics such as fibrin fibre diameter and fibre density, as determined by scanning electron microscopic image analysis. The maximum amplitude (MA) as measured by TEG, which purports to relate to clot strength, was unable to detect these differences

Measurement of total cholesterol using an enzyme sensor based on a printed hydrogen peroxide electrocatalyst

© 2016 The Royal Society of Chemistry. Cholesterol is a major modifiable risk factor in cardiovascular disease and can be effectively managed by a combination of medication and monitoring. There continues to be a need for new point-of-care diagnostics to measure lipid panels, including total cholesterol. Enzyme assays based on the generation of hydrogen peroxide have been very effective in this regard. This work demonstrates the application of printed electrochemical sensors to the measurement of total cholesterol in serum. The assay uses the surfactant Triton X-100 to provide electrocatalytic enhancement of a silver paste screen-printed electrode to hydrogen peroxide, while also achieving effective solubilisation of total cholesterol from lipoprotein. The resulting biosensors employed 0.5% (v/v) Triton X-100 in PBS with 156 U mL-1 cholesterol esterase and 60 U mL-1 cholesterol oxidase. Measurement of total cholesterol in serum in the range of 0 to 10 mM had a sensitivity of 2.24 × 10-8 A mM-1, with coefficient of determination of 0.984, detection limit of at least 2 mM and average relative standard deviation of 10.8% (n = 3)