Amperometric Screen-Printed Galactose Biosensor for Cell Toxicity Applications

© 2016, Copyright © Taylor & Francis Group, LLC. ABSTRACT: This paper reports the development and application of a biosensor for the amperometric determination of galactose in the presence of human hepatocellular carcinoma cells with and without a hepatotoxic agent. The biosensor was fabricated by drop-coating 1.5% cellulose acetate on a 3×3mm screen-printed carbon electrode followed by depositing 2 U of galactose oxidase. The electrodes dimensions were reduced to 3×0.5mm before measurements. Hepatocellular carcinoma cells were utilized for in vitro toxicity testing by evaluating the effect of paracetamol on galactose uptake. The amperometric responses to galactose indicated that the inhibition of uptake was directly proportional to the concentration of paracetamol following 24h of exposure to the hepatocellular carcinoma cells. These results demonstrate that the fabricated biosensor may be used for the real-time monitoring of cell metabolism and toxicity

Homogeneous functional self-assembled monolayers: Faradaic impedance baseline signal drift suppression for high-sensitivity immunosensing of C-reactive protein

The limit of detection (LOD) of affinity sensors based on alkanethiol self-assembled monolayers (SAMs) system can be improved by either signal amplification and/or noise reduction. The latter includes baseline signal drift arising, in part, from, monolayer imperfections (and variations in this through repeats) as well as electrical noise of both the sensor and transducer. In reagentless “label free” assays signal drift carries with it the possibility of assay false-positive results (if signal drift is positive) or false-negatives (if specific signal is swamped by downward drift). Faradaic electrochemical impedance spectroscopy (FEIS) sensors based on SAM interfaces have been shown to exhibit considerable baseline signal drift, which consequently affects their assaying capabilities. This study reports on the development of a simple two-step pre-treatment method for generating functional SAMs of 11-mercaptoundecanoic acid (MUA) on Au with a highly suppressed baseline signal drift. For electrochemical characterisation of the SAMs, the charge-transfer resistance (Rct), defect presence (pinhole radius and separation), capacitance, and dielectric constant were evaluated. The interface pre-treatment generates films of substantially improved homogeneity that are subsequently functionalised with anti-human C-reactive protein. CRP, an acute-phase protein, is detectable down to femtomolar levels without any amplification; this is a 2–3 order of magnitude lower detection limit than that typically accessible for analyses of this type. The developed protocols thus present a convenient and general route to suppress baseline signal drifts and improve the limits of detection of affinity sensors based on Faradaic impedance

A Voltammetric Sensor Based on Chemically Reduced Graphene Oxide-Modified Screen-Printed Carbon Electrode for the Simultaneous Analysis of Uric Acid, Ascorbic Acid and Dopamine

A disposable screen-printed carbon electrode (SPCE) modified with chemically reduced graphene oxide (rGO) (rGO-SPCE) is described. The rGO-SPCE was characterized by UV-Vis and electrochemical impedance spectroscopy, and cyclic voltammetry. The electrode displays excellent electrocatalytic activity towards uric acid (UA), ascorbic acid (AA) and dopamine (DA). Three resolved voltammetric peaks (at 183 mV for UA, 273 mV for AA and 317 mV for DA, all vs. Ag/AgCl) were found. Differential pulse voltammetry was used to simultaneously detect UA, AA and DA in their ternary mixtures. The linear working range extends from 10 to 3000 μM for UA; 0.1 to 2.5 μM, and 5.0 to 2 × 104 µM for AA; and 0.2 to 80.0 μM and 120.0 to 500 µM for DA, and the limits of detection (S/N = 3) are 0.1, 50.0, and 0.4 μM, respectively. The performance of the sensor was evaluated by analysing spiked human urine samples, and the recoveries were found to be well over 98.0% for the three compounds. These results indicate that the rGO-SPCE represents a sensitive analytical sensing tool for simultaneous analysis of UA, AA and DA

Immunochemical Assays and Nucleic-Acid Detection Techniques for Clinical Diagnosis of Prostate Cancer

Prostate cancer (PCa) is a significant cause of morbidity and mortality and the most common cancer in men in Europe, North America, and some parts of Africa. The established methods for detecting PCa are normally based on tests using Prostate Specific Antigen (PSA) in blood, Prostate cancer antigen 3 (PCA3) in urine and tissue Alpha-methylacyl-CoA racemase (AMACR) as tumour markers in patient samples. Prior to the introduction of PSA in clinics, prostatic acid phosphatase (PAP) was the most widely used biomarker. An early diagnosis of PCa through the detection of these biomarkers requires the availability of simple, reliable, cost-effective and robust techniques. Immunoassays and nucleic acid detection techniques have experienced unprecedented growth in recent years and seem to be the most promising analytical tools. This growth has been driven in part by the surge in demand for near-patient-testing systems in clinical diagnosis. This article reviews immunochemical assays, and nucleic-acid detection techniques that have been used to clinically diagnose PCa

Ultrasensitive impedimetric immunosensor for the detection of C-reactive protein in blood at surface-initiated-reversible addition–fragmentation chain transfer generated poly(2-hydroxyethyl methacrylate) brushes

The reversible addition–fragmentation chain transfer (RAFT) polymerization of 2-hydroxethyl methacrylate (HEMA) from a surface confined, dithio-tethered, chain transfer agent (CTA) enables the preparation of electrode-tethered poly(2-hydroxyethyl methacrylate) (pHEMA) brushes of well-defined thickness with convenience and exceptionally high interfacial impedimetric baseline stability. The subsequent covalent integration of antibodies generates interfaces of very high target recognition specificity, ultimately enabling femtomolar levels of quantification of C-reactive protein (CRP) and recovery in spiked serum samples of ∼98%. When combined with the intrinsic scalability of the reagentless electrochemical impedance spectroscopy (EIS) platform, and the innate high levels of polymer tuneability and control, we believe this represents a valuable contribution to the diagnostic toolbox

Functional molecular interfaces for impedance-based diagnostics

In seeking to develop and optimize reagentless electroanalytical assays, a consideration of the transducing interface features lies key to any subsequent sensitivity and selectivity. This review briefly summarizes some of the most commonly used receptive interfaces that have been employed within the development of impedimetric molecular sensors. We discuss the use of high surface area carbon, nanoparticles, and a range of bioreceptors that can subsequently be integrated. The review spans the most commonly utilized biorecognition elements, such as antibodies, antibody fragments, aptamers, and nucleic acids, and touches on some novel emerging alternatives such as nanofragments, molecularly imprinted polymers, and bacteriophages. Reference is made to the immobilization chemistries available along with a consideration of both optimal packing density and recognition probe orientation. We also discuss assay-relevant mechanistic details and applications in real sample analysis