Ihab Abdel-Hamid

Biosensors for detection of pathogenic bacteria

Abstract This paper presents an overview of different physicochemical instrumental techniques for direct and indirect identification of bacteria such as: infrared and fluorescence spectroscopy, flow cytometry, chromatography and chemiluminescence techniques as a basis for biosensor construction. A discussion of publications dealing with emerging biosensors for bacterial detection is presented. The review presents recent advances in the development of alternative enzyme- and immunosensors for detection of pathogenic bacteria in a variety of fields (e.g. clinical diagnostics, food analysis and environmental monitoring). Depending on the biological element employed: enzyme; nucleic acid and antibody based biosensors are discussed. Depending on the basic transducer principles, recent advances in biosensing technologies that use electrochemical, piezoelectric, optical, acoustic and thermal biosensors for detection of pathogenic bacteria are overviewed. Special attention is paid to methods for improving the analytical parameters of biosensors including sensitivity and analysis time as well as automation of assay procedures. Recent developments in immunofiltration, flow-injection and flow-through biosensors for bacterial detection are overviewed from the system’s engineering point of view. Future directions for biosensor development and problems related to the commercialization of bacterial biosensors are discussed in the final part of this review.

Application of Electrochemical Biosensors for Detection of Food Pathogenic Bacteria

Current practices for preventing microbial diseases rely upon careful control of various kinds of pathogenic bacteria in food safety and environmental monitoring. The main disadvantages of conventional bacterial detection methods are the multistep procedure and long time requirements. This article gives an overview of alternative electrochemical biosensors for detection of pathogenic bacteria in the food industry. Focus has been on new microbial metabolism-based, antibody-based and DNA-based biosensors. The underlying principles and applications of these biosensors are discussed. Recent developments in flow-injection biosensor systems with an electrochemical detection are also presented.

Flow-through immunofiltration assay system for rapid detection of E. coli O157 :H7

A flow-through amperometric immunofiltration assay system based on disposable porous filter-membranes for rapid detection of Escherichia coli O157:H7 has been developed. The analytical system utilizes flow-through, immunofiltration and enzyme immunoassay techniques in conjunction with an amperometric sensor. The parameters affecting the immunoassay such as selection of appropriate filter membranes, membrane pore size, antibody binding capacity and the concentrations of immunoreagents were investigated and optimized. Non-specific adsorption of the enzyme conjugate was investigated and minimized. A sandwich scheme of immunoassay was employed and the immunofiltration system allows to specifically and directly detect E. coli cells with a lower detection limit of 100 cells/ml. The working range is from 100 to 600 cells/ml with an overall analysis time of 30 min. No pre-enrichment was needed. This immunosensor can be easily adapted for assay of other microorganisms and may be a basis for a new class of highly sensitive bioanalytical devices for rapid quantitative detection of bacteria.

Highly sensitive flow-injection immunoassay system for rapid detection of bacteria☆

A flow-injection amperometric immunofiltration assay system for the rapid detection of total Escherichia coli and Salmonella was developed. The system is based on the use of disposable porous nylon membranes which act as a support for the immobilization of anti-E. coli or anti-Salmonella antibodies. The assay system consists of a flow-injection system, a disposable filter-membrane and an amperometric sensor. Parameters affecting the performance of the immunofiltration assay system such as membrane pore size, non-specific binding, conjugate concentration and sample volume were studied and optimized. A sandwich scheme of immunoassay was employed and the immunofiltration system was able to specifically and directly detect 50 cells/ml of total E. coli or 50 cells/ml of Salmonella with an overall analysis time of 35 min. This immunosensor can be easily adapted for the assay of other microorganisms and may be a basis for a new class of highly sensitive and automated bioanalytical devices for the rapid quantitative detection of bacteria.

DEVELOPMENT OF A FLOW-THROUGH IMMUNOASSAY SYSTEM

Abstract A semi-automated flow-through immunoassay system consisting of an amperometric immunosensor and reagent flow arrangements has been developed. The amperometric immunosensor is based on a high-surface-area carbon immunoelectrode. The novel principle of flow immunoelectrode based on highly dispersed carbon material which acts as both an immunosorbent and an immunoelectrode was employed. The immunoelectrode is designed to be a disposable sensing element. The ‘sandwich’ scheme of immunoassay has been used and iodine formed as a result of the enzymatic oxidation of iodide by peroxidase-label has been detected amperometrically. The overall time of analysis including flowing of analyte, flowing of antigen, washing and detecting stages is as low as 17 min. The developed system allows fast determination of rabbit IgG (used as a model analyte) with a low detection limit in the 10−11 M range and may easily be adapted for detection of various other analyses. The immunoassay system has the potential for miniaturization and complete automation allowing its use in laboratory setting as well as in field conditions.

Fast Amperometric Assay for E. coli O157:H7 Using Partially Immersed Immunoelectrodes

A novel amperometric immunoelectrode for fast and sensitive assay of E. coli O157:H7 is presented. Antibodies against E. coli O157:H7 were immobilized on the surface of carbon rods, which acted as both working electrode and sorbent surface. A sandwich scheme of immunoassay was used and 5-aminosalicylic acid was employed as a redox mediator for the amperometric detection of the enzyme-label (horseradish peroxidase). The immunoelectrodes were operated while being partly immersed in the detection cell which resulted in the acceleration of the diffusion-controlled rates of immunological, enzymatic and electrochemical reactions. The amperometric immunoelectrode allows the achievement of significantly lower detection limits (40 times lower) than that achievable with standard spectrophotometric detection ELISA method using the same immunochemicals. The immunoelectrode allows determination of E. coli cell concentrations in the range from 200 to 7000 cells/mL with an overall analysis time of 40 min. This immunoelectrode can be easily adapted for assay of other microorganisms and may be a basis for creating a new class of highly sensitive and rapid immunosensors.

Development of a needle-type biosensor for intravascular glucose monitoring☆

Abstract A needle type glucose biosensor has been developed for monitoring glucose levels during the hemorrhagic shock in trauma patients. The sensor employs polyurethane, cellulose acetate or PVC layers as an outer glucose diffusion and protective membrane, and glucose oxidase enzyme entrapped in poly(1,3-phenylenediamine) film. Sensor performance was evaluated in vitro and the sensor shows a sensitivity of up to 35 nA mM and a linear range of up to 38 mM. Evaluation of the sensor response in serum showed similar sensitivity and linear range as obtained using calibration curves in buffer solution. The sensor has a short response time of 23 s. The sensors were operated continuously for 14 days in phosphate buffer solution, and no significant change in the sensitivity and the linear range was observed during the first 5 days. Sensors show a minimum change in their performance when stored inactive in buffer at 4 °C for at least eight weeks.

Development of a needle type glucose biosensor

Abstract A needle type glucose biosensor has been developed for monitoring of glucose levels during hemorrhagic shock. The sensor employs polycarbonate membranes that are either untreated or treated with Nafion or Silastic, and Glucose Oxidase enzyme which is immobilized on modified carbon powder in a gel matrix. Sensor performance was evaluated in vitro and the sensor showed a sensitivity of up to 10 nA/mM and a linear range of up to 30 mM. Evaluation of the sensor response in serum showed similar sensitivity and linear range as obtained using calibration curves in buffer solution. The sensors were operated continuously for 28 days in phosphate buffer solution and no significant change in the sensitivity and the linear range was observed during the first 7 days. Sensors show a minimum change in their performance when stored inactive in buffer at 4°C for at least eight weeks.