Characterization of an Electroless Plated Silver/Cysteine Sensor Platform for the Electrochemical Determination of Aflatoxin B 1 (original) (raw)
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An electroless plated silver/cysteine sensor platform [Glass|silver|cysteine|aflatoxin B1|horseradish peroxidase] for the Electrochemical detection of aflatoxin B1 was developed and characterized. This involved four major steps: (1) an electroless deposition of silver (plating) onto a glass slide, (2) immobilization of cysteine; (3) conjugation of aflatoxin B1 to cysteine groups; and (4) blocking of free cysteine groups with horseradish peroxidase (HRP).The binding of cysteine to the silver was demonstrated by the disappearance of thiol (S-H) groups at 2500 cm−1 using Fourier transmittance infrared spectra (FT-IR), while the subsequent steps in the assembly of sensor platform were monitored using both FT-IR and cyclic voltammetry, respectively. The sensor platform exhibited a broadened nonsymmetrical redox couple as indicated by cyclic voltammetry. The platform was further characterized for sensitivity and limit of detection. The indirect competitive immunoassay format, whereby free and immobilized aflatoxin B1 on the sensor competed for the binding site of free anti-aflatoxin B1 antibody, was used at various concentrations of aflatoxin B1. The sensor generated differential staircase voltammogram that was inversely proportional to the concentration of aflatoxin B1 and aflatoxin B1 in the range of 0.06–1.1 ng/mL with a detection limit of 0.08 ng/mL could be detected.
The development of a field-deployable biosensor device for aflatoxin detection is an attractive endeavor that should solve current bottlenecks related to the current chromatographic and spectroscopic techniques including: the requirement for highly trained operators, time consuming between analyses and obtaining of results, the requirement for labeling in some instances, and high costs of the equipment, among others. Besides, such an analytical device should: requires smaller sample volumes, release of analytical results fast, cheap, precise as well as accurate, and used at the point of need. To achieve these goals, the newly developed biosensor platform (electroless plated silver/cysteine sensor platform [Glass|silver|cysteine|aflatoxin B1|horseradish peroxidase]; Wacoo et al, [1], was functionalized into an electrochemical aflatoxin B1 biosensor device. The biosensor prototype was assembled by interfacing: (1) the sensor platform containing biorecognition element and the traducer with (2) electronic relay system made of potentiostat with PIC16F877A microcontroller, the liquid-crystal display (LCD), and the system powered by a replaceable 5 volt battery. The immunosensor prototype successfully generated a quasi-reversible redox peak characteristic of the developed sensor platform at 600 mV and a differential staircase voltammogram (DSCV) cathodic peak at 304 mV with the peak height of 5920 mV. Therefore, when this prototype is validated for aflatoxin B1 analysis, it could be used for analysis of aflatoxin B1 in food samples.
Impedimetric aflatoxin M1 immunosensor based on colloidal gold and silver electrodeposition
Sensors and Actuators B: Chemical, 2009
An impedimetric immunosensor based on colloidal gold and silver electrodeposition for the detection of aflatoxin M 1 (AFM 1) herein is reported. An indirect like competitive ELISA procedure was performed on screen-printed electrodes (SPEs) in presence of anti-AFM 1 gold-labelled antibodies. Silver was chronoamperometrically electrodeposited at a fixed applied potential and for a determined period of time to amplify the signal. The calculated charge transfer resistance (R ct) was found to correlate well with the concentration of AFM 1. The linear working range of the described AFM 1 immunosensor ranged between 15 and 1000 ng/L with a limit of detection (LoD) equal to 15 ng/L (R.S.D. = 20%). Impedimetric results were confronted with linear sweep voltametry (LSV) and corresponded well to this technique.
Recent Advances in Electrochemical-Based Sensing Platforms for Aflatoxins Detection
Chemosensors, 2016
Mycotoxin are small (MW~700 Da), toxic secondary metabolites produced by fungal species that readily colonize crops and contaminate them at both pre-and post-harvesting. Among all, aflatoxins (AFs) are mycotoxins of major significance due to their presence in common food commodities and the potential threat to human health worldwide. Based on the severity of illness and increased incidences of AFs poisoning, a broad range of conventional and analytical detection techniques that could be useful and practical have already been reported. However, due to the variety of structural analogous of these toxins, it is impossible to use one common technique for their analysis. Numerous recent research efforts have been directed to explore alternative detection technologies. Recently, immunosensors and aptasensors have gained promising potential in the area of sample preparation and detection systems. These sensors offer the advantages of disposability, portability, miniaturization, and on-site analysis. In a typical design of an aptasensor, an aptamer (ssDNA or RNA) is used as a bio-recognition element either integrated within or in intimate association with the transducer surface. This review paper is focused on the recent advances in electrochemical immunoand aptasensing platforms for detection of AFs in real samples.
2008
An aflatoxin B 1 (AFB 1 ) electrochemical immunosensor was developed by the immobilisation of aflatoxin B 1 -bovine serum albumin (AFB 1 -BSA) conjugate on a polythionine (PTH)/gold nanoparticles (AuNP)-modified glassy carbon electrode (GCE). The surface of the AFB 1 -BSA conjugate was covered with horseradish peroxidase (HRP), in order to prevent non-specific binding of the immunosensors with ions in the test solution. The AFB 1 immunosensor exhibited a quasi-reversible electrochemistry as indicated by a cyclic voltammetric (CV) peak separation (ΔE p ) value of 62 mV. The experimental procedure for the detection of AFB 1 involved the setting up of a competition between free AFB 1 and the immobilised AFB 1 -BSA conjugate for the binding sites of free anti-aflatoxin B 1 (anti-AFB 1 ) antibody. The immunosensor's differential pulse voltammetry (DPV) responses (peak currents) decreased as the concentration of free AFB 1 increased within a dynamic linear range (DLR) of 0.6 -2.4 ng/mL AFB 1 and a limit of detection (LOD) of 0.07 ng/mL AFB 1 . This immunosensing procedure eliminates the need for enzyme-labeled secondary antibodies normally used in conventional ELISA-based immunosensors.
Because of the potential health impact of aflatoxin B 1 (AFB 1 ), it is essential to monitor the level of this mycotoxin in a variety of foods and agricultural products. In this paper, a novel immunosensor for the rapid detection of AFB 1 based on label-free electrochemical impedance spectroscopy (EIS) monitoring was achieved. The immunosensor was fabricated by stepwise immobilization of 1,6-hexanedithiol, colloidal Au, and aflatoxin B 1 -bovine serum albumin conjugate (AFB 1 -BSA) on a gold electrode via selfassembling technique. The interfacial properties of the modified electrodes were evaluated using the Fe(CN) 6 3À/4À redox couple as a probe via cyclic voltammetry (CV) and EIS. An equivalent circuit model with a constant phase element was used to interpret the obtained impedance spectra. The impedance via the specific immuno-interaction at the sensor surface was utilized to detect AFB 1 in samples. Under the optimized conditions, the impedance increment was linearly related to the AFB 1 concentration in the range of 0.08 to 100 ng mL À1 with a detection limit of 0.05 ng mL À1 (S/N ¼ 3) and a correlation coefficient of 0.9919.
Because of the potential health impact of aflatoxin B 1 (AFB 1 ), it is essential to monitor the level of this mycotoxin in a variety of foods and agricultural products. In this paper, a novel immunosensor for the rapid detection of AFB 1 based on label-free electrochemical impedance spectroscopy (EIS) monitoring was achieved. The immunosensor was fabricated by stepwise immobilization of 1,6-hexanedithiol, colloidal Au, and aflatoxin B 1 -bovine serum albumin conjugate (AFB 1 -BSA) on a gold electrode via selfassembling technique. The interfacial properties of the modified electrodes were evaluated using the Fe(CN) 6 3À/4À redox couple as a probe via cyclic voltammetry (CV) and EIS. An equivalent circuit model with a constant phase element was used to interpret the obtained impedance spectra. The impedance via the specific immuno-interaction at the sensor surface was utilized to detect AFB 1 in samples. Under the optimized conditions, the impedance increment was linearly related to the AFB 1 concentration in the range of 0.08 to 100 ng mL À1 with a detection limit of 0.05 ng mL À1 (S/N ¼ 3) and a correlation coefficient of 0.9919.
Electrochemical Immunochip Sensor for Aflatoxin M1 Detection
Analytical Chemistry, 2009
An investigation into the fabrication, electrochemical characterization, and development of a microelectrode array (MEA) immunosensor for aflatoxin M 1 is presented in this paper. Gold MEAs (consisting of 35 microsquare electrodes with 20 µm × 20 µm dimensions and edge-to-edge spacing of 200 µm) together with onchip reference and counter electrodes were fabricated using standard photolithographic methods. The MEAs were then characterized by cyclic voltammetry, and the behavior of the on-chip electrodes were evaluated. The microarray sensors were assessed for their applicability to the development of an immunosensor for the analysis of aflatoxin M 1 directly in milk samples. Following the sensor surface silanization, antibodies were immobilized by cross-linking with 1,4-phenylene diisothiocyanate (PDITC). Surface characterization was conducted by electrochemistry, fluorescence microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM). A competitive enzyme linked immunosorbent assay (ELISA) assay format was developed on the microarray electrode surface using the 3,3,5′,5′-tetramethylbenzidine dihyrochloride (TMB)/ H 2 O 2 electrochemical detection scheme with horseradish peroxidase (HRP) as the enzyme label. The performance of the assay and the microarray sensor were characterized in pure buffer conditions before applying to the milk samples. With the use of this approach, the detection limit for aflatoxin M 1 in milk was estimated to be 8 ng L-1 , with a dynamic detection range of 10-100 ng L-1 , which meets present legislative limits of 50 ng L-1. The milk interference with the sensor surface was also found to be minimal. These devices show high potential for development of a range of new applications which have previously only been detected using elaborate instrumentation.
Biosensors & Bioelectronics, 2005
The production and assembling of disposable electrochemical AFM1 immunosensors, which can combine the high selectivity of immunoanalysis with the ease of the electrochemical probes, has been carried out. Firstly immunoassay parameters such as amounts of antibody and labelled antigen, buffer and pH, length of time and temperature of each steps (precoating, coating, binding and competition steps) were evaluated and optimised in order to set up a spectrophotometric enzyme-linked immunosorbent assay (ELISA) procedure. This assay exhibited a working range between 30 and 160 ppt in a direct competitive format. Then electrochemical immunosensors were fabricated by immobilising the antibodies directly on the surface of screen-printed electrodes (SPEs), and allowing the competition to occur between free AFM1 and that conjugated with peroxidase (HRP) enzyme. The electrochemical technique chosen was the chronoamperometry, performed at −100 mV. Furthermore, studies of interference and matrix effects have been performed to evaluate the suitability of the developed immunosensors for the analysis of aflatoxin M1 directly in milk. Results have shown that using screen-printed electrodes aflatoxin M1 can be measured with a detection limit of 25 ppt and with a working range between 30 and 160 ppt. A comparison between the spectrophotometric and electrochemical procedure showed that a better detection limit and shorter analysis time could be achieved using electrochemical detection.