Immunosensors types

A novel immunosensor type for low-molecular weight analytes has... 

This immunosensor-type assay uses ferrocene methanol as co-substrate (redox mediator) of glucose oxidase (GOD). The various steps of the assay, shown in Fig. 8.20, are as follows (1) Covalent immobilization of protein A on graphite-polystyrene screen-printed electrodes (SPEs) (2) Addition of the rabbit IgG to be quantified which is captured specifically by protein A (3) Addition of a biotinylated goat anti-rabbit antibody (4) Addition of avidin-GOD conjugate (5) Addition of glucose and ferrocene methanol (6) Measurement of catalytic current by flow injection immunoassay. The voltammetric current corresponds to the one-electron oxidation of the ferrocenyl group to ferricinium. The electrode can subsequently be regenerated up to 30 times. The feasibility of the assay has been demonstrated for the case of monoclonal mouse anti-human prolactin (PL) with a detection limit of 0.02 pg mL [90]. 

Enzyme Immunosensors. Enzyme immunosensors are enzyme immunoassays coupled with electrochemical sensors. These sensors (qv) require multiple steps for analyte determination, and either sandwich assays or competitive binding assays maybe used. Both of these assays use antibodies for the analyte of interest attached to a membrane on the surface of an electrochemical sensor. In the sandwich assay type, the membrane-bound antibody binds the sample antigen, which in turn binds another antibody that is enzyme-labeled. This immunosensor is then placed in a solution containing the substrate for the labeling enzyme and the rate of product formation is measured electrochemically. The rate of the reaction is proportional to the amount of bound enzyme and thus to the amount of the analyte antigen. The sandwich assay can be used only with antigens capable of binding two different antibodies simultaneously (53). 

There are mainly three types of transducers used in immunosensors electrochemical, optical, and microgravimetric transducers. The immunosensors may operate either as direct immunosensors or as indirect ones. For direct immunosensors, the transducers directly detect the physical or chemical effects resulting from the immunocomplex formation at the interfaces, with no additional labels used. The direct immunosensors detect the analytes in real time. For indirect immunosensors, one or multiple labeled bio-reagents are commonly used during the detection processes, and the transducers should detect the signals from the labels. These indirect detections used to need several washing and separation steps and are sometimes called immunoassays. Compared with the direct immunosensors, the indirect immunosensors may have higher sensitivity and better ability to defend interference from non-specific adsorption. 

Moreover, there is another important type of microgravimetric immunosensor which is based on the immunological agglutination events. The agglutination immunoreaction... 

Immunosensors have been developed commercially mostly for medical purposes but would appear to have considerable potential for food analysis. The Pharmacia company has developed an optical biosensor, which is a fully automated continuous-flow system which exploits the phenomenon of surface plasmon resonance (SPR) to detect and measure biomolecular interactions. The technique has been validated for determination of folic acid and biotin in fortified foods (Indyk, 2000 Bostrom and Lindeberg, 2000), and more recently for vitamin Bi2. This type of technique has great potential for application to a wide range of food additives but its advance will be linked to the availability of specific antibodies or other receptors for the various additives. It should be possible to analyse a whole range of additives by multi-channel continuous flow systems with further miniaturisation. 

Electrochemical immunosensors are a powerful tool for the analysis of antibacterials in food and different configurations have been presented during recent years. For example, an amperometric immunosensor was reported by Wu et al. [182], for penicillin quantification in milk, with a linear range from 0.25 to 3 ng/ml and a limit of detection of 0.3 pg/L [182]. Other types of transduction have been also explored, like a label-free impedimetric flow injection immunosensor for the detection of penicillin G. 

Aizawa et al. developed immunosensors of the first type for syphilis and blood typing [228-230] based on measurements of the transmembrane potential across an immunoresponsive membrane. The potentiometric immunosensor using an antibody against human chorionic gonadotropin (hCG)... 

Types of biosensors can be named either by the biological components, physical transducing devices, or the measured analytes. Researchers were originally using biological components to define types of biosensors (Table 2). Types of transducers had also been included in the name to identify the physical transducing device, i.e., enzyme electrodes, acoustic-immunosensors, optical biosensors, piezoelectric-immunosensors, and biochips. 

Very few immunosensors are commercially available. The commercial immunosensors are either the detector or bioanalyzer types. The PZ 106 immunosensor from Universal Sensors Inc. (New Orleans, LA) has been used as a detector to measure antibody-antigen reaction. Ohmicron (Newtown, PA) developed a series of pesticide immuno-bioanalyzers that have been used in field tests. Pharmacia Biosensor USA (Piscataway, NJ) recently introduced BIAcore immunodetection system. A combination of a unique flow injection device and surface plasmon resonance (SPR) detection technique provides a real time analysis. A carboxylmethyldextran layer added to plasmon generating gold film is a hydrophobic, activatable, and flexible polymer that provides high antibody and low non-specific bindings. System demonstration at the Institute of Food Technologists (IFT) 1994 meeting in Atlanta drew attention of food scientists. It should easily be adapted for food protein characterization. 

Haga et al. developed another type of immunosensor by combining an enzyme membrane immunoassay and an enzyme sensor using oxygen electrodes (HI). In this assay antigen molecules (theophylline) are attached on the surface of the liposomes and an enzyme (horseradish peroxidase) is encapsulated in the sensitized liposome. When antibody (antitheophylline antibody) and complement are added, the enzyme is released by the liposome lysis. The enzyme activity with the NADH-NAD reaction can be determined by the oxygen electrode. When antigen is added, it competitively binds to antibodies, then liposome lysis and enzyme activity are decreased. The sensitivity of this method for theophylline determination was reported as 0.7 ng/ml. 

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