Continuous flow micro-cell for electrochemical addressing of engineered bio-molecules

Abstract A micro-cell consisting of a planar array of Au working electrodes, covered by a glass reservoir (600 nl capacity), connected with perpendicular capillaries and provided with an Ag/AgCl wire reference electrode (RE) has been manufactured. Addressed on-chip immobilization and sensing of biomolecules was demonstrated by the immobilization of histidine (HIS) tagged alkaline phosphatase (AP) under continuous flow (10 l/min) conditions on the base of electrochemically deposited multilayer (EDM) of cysteamine modified by nitrilotriacetic acid (NTA). The presented method allows the immobilization of different biomolecules on the ready-to-use reservoir covered chip without needs of manual biomolecule deposition. Further on, the detection of analytes can be performed using the electrochemical measurements. Since the methods of biomolecule deposition and sensing are both electrochemical, this allows simplifying the production technology using cheaper detection instruments compared to prevailing present-day optical detection modes

Immobilisation of engineered molecules on electrodes and optical surfaces,

ABSTRACT Monolayers of genetically modified proteins with an hexahistidine tag, (His)6, were obtained by using a Ni -NTA chelator synthesized on gold sputtered surfaces (via sulphide bonds), or on gold and graphite (via sililating agents) working electrodes of screen-printed devices. Two kinds of proteins were produced and purified for this study: a) a recombinant antibody, derived from the 'single chain Fv' ( scFv) format; b) a photosystem II (PSII) core complex isolated from the mutant strain CP43-H of the thermophilic cyanobacterium Synechococcus elongatus. An scFv, previously isolated from a synthetic 'phage display' library, was further engineered with an alkaline phosphatase activity genetically added between the carbossi-terminal of the scFvs and the (His)6 to allow direct measurement of immobilisation. Renewable specific binding of (His)6-proteins to gold and graphite surfaces and fast and sensitive electrochemical or optical detection of analytes were obtained. Additionally, "on chip" protein preconcentration was conveniently achieved for biosensing purposes, starting from crude unpurified extracts and avoiding protein purification steps

Disposable label - Free immunosensor on screen - Printed electrodes for the detection of Aflatoxin B1 in food

An impedimetric label - free immunosensor on disposable screen-printed carbon electrode (SPCE) for Aflatoxin B1 detection has been developed in this work. SPCE surface was modified with gold nanoparticles and the monoclonal anti-AFB1 was immobilized on the working electrode through a cysteamine layer. After each coating step, the modified surface was characterized by Cyclic Voltammetry and Electrochemical Impedance Spectroscopy. The charge transfer resistance (Rct) was chosen as the best parameter that describes the reproducible change in electrical properties of the electrode surface at different AFB1 concentrations and it was used to investigate the analytical parameters of the developed immunosensor. Under optimized conditions, the immunosensor showed a linear relationship between 0.01 and 4 ng/mL with a low detection limit of 0.01 ng/mL, making it suitable to detect AFB1 amount in many common food products. Lastly, the immunosensor was used to measure AFB1 in nuts samples with good results

Impedimetric label-free immunosensor for rapid detection of Ochratoxin A in beer and wine

In this work, the development of an impedimetric label-free immunosensor for the detection of Ochratoxin A in food products is reported. Two different immobilization schemes of a monoclonal antibody for OTA detection were investigated and compared, with the aim to develop an analytical device able to detect an OTA amount equal to the maximum levels imposed by European legislation for food products. activated ferrocene as electron-transferring mediator, improved the electrical properties of the immunosensor, which showed a limit of detection equal to 0.25 ng/mL. This immunosensor was used to analyze wine and beer samples spiked with known OTA amounts

Entirely automated glucose monitoring system based on a flow injection analysis apparatus

A process for waste whey treatment, fully based on membrane technologies, has been studied and developed in a pilot plant scale, with the aim to recover chemicals from farm wastes (fats, whey proteins, lactose, salts and water). This process, described in its essential steps, includes a hollow fiber membrane reactor for the hydrolysis of lactose with an immobilized β-galactosidase. The bioreactor performances are studied, in a lab scale, with an entirely automated glucose monitoring system based on a flow injection analysis apparatus. The monitoring system shows interesting performances concerning signal stability, extended linearity (up to 150 mmol l-1) and long term monitoring (up to 3 months without manual operations). The performances of the glucose monitoring system are described. © 1994, Taylor & Francis Group, LLC. All rights reserved

Label-free impedimetric immunosensors for sensitive detection of aflatoxin B1 in food

Aflatoxin B1 (AFB1) is one of the mycotoxins highly resistant to food processing that enters the food chain and provide a threat to human health. In this paper, different immobilization techniques of the monoclonal anti-AFB1 on gold electrodes were tested and compared, in order to develop a label-free immunosensor able to detect AFB1 amount imposed by European legislation for adult and infant foods. Different materials, coupled with Electrochemical Impedance Spectroscopy as transduction technique, were used to improve immunosensors analytical parameters, such as linear range, sensitivity and limit of detection. Through the use of ferrocene molecules, the immunosensor showed linearity in the range 0.01-10 ng/mL and the lowest limit of detection (0.01 ng/mL), allowing the possibility to use it in a wide range of food products, including infant foods

Label-free impedimetric biosensors for the control of food safety–a review

The guarantee of food safety requires fast and accurate control for all contaminants, chemicals and bacteria, which are harmful to human health. Ensuring the safety and quality of food is the main interest both for the food industry and for consumers. In the food industry, the safety of a food product is evaluated through chemical and microbiological analysis: these procedures conventionally use the technique as chromatography, spectrophotometry and electrophoresis that are time-consuming, require highly trained personnel, are expensive and require steps of sample pretreatment, increasing the time of analysis. Consequently, among food and beverage industries, exists a growing demand in biosensing technologies as simple, rapid, accurate, low - cost and portable analytical devices for the monitoring and detection of chemical and microbiological contaminants (toxins, mycotoxins, pathogenic bacteria, pesticides and allergens) that endanger the food safety. Among biosensors, the label-free biosensors, characterised by direct detection of the analyte of interest, exploiting the advantages of an impedimetric transduction technique, seem to be the most promising devices for the future. In the last years, the potential use of Electrochemical Impedance Spectroscopy technique has been widely considered in the biosensor field; it is, in fact, a powerful, non-destructive and informative technique which can be used to study the electrical properties of the sensing device interface and to trace the reactions that occur on it. For these reasons, the application of impedimetric technique as a transduction technology has allowed the label-free detection and sensitive quantification of the bio interaction in different application and thus for the development of food hazards biosensors too. This review provides the current situation in the literature on label-free impedimetric biosensors in food safety