Immunosensors biotin

FIGURE 5.5 Schematic representations of the two immunosensor formats (a) immunosensor based on the biotin-streptavidin interaction and (b) immunosensor based on rabbit IgG-modified SPCEs. (Reprinted from [27] with permission from Elsevier.)... 

O. Ouerghi, A. Touhami, N. Jaffrezic-Renault, C. Martelet, H. Ben Ouada, and S. Cosnier, Impedimetric immunosensor using avidin-biotin for antibody immobilization. Bioelectrochemistry 56, 131—133... 

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. 

Liu et al. [140] have also used this interface for an electrochemical immunosensor for small molecules (Figure 1.26). In this sensor, one end of the molecular wire is attached to ferrocene dimethylamine with a covalent link formed between one of the amine group son the ferrocene and the carboxyl group on the wire. To the other amine is attached the antibody-binding epitope for the antibody, in this proof-of-concept study the epitope is biotin. Electron transfer can be readily achieved to the ferrocene molecule but upon antibody binding to this interface, the electrochemical signal is dramatically reduced. 

One of the most important features in the immunosensor design is the proper choice of the immobilization method for keeping the affinity of the antibodies. As was previously demonstrated for Protein A, when the antibodies are immobilized through their Fc fragment to Protein A (or G), their Fab binding sites are mostly oriented away from the solid phase. As Protein A is able to link the Fc region of different antibodies, there is no need to modify the antibody with biotin. As an antecedent, we have previously demonstrated the utility of Protein A biocomposite (ProtA-GEB) for the universal attachment of antibodies with different specificities [54]. 

Fig. 26.2. Amperometric immunosensors set-up using a biotinylated copolymer poly(pyrrole-biotin, pyrrole-lactitob-ionamide) coated platinum or glassy carbon electrodes and three enzymatic markers (GOX-B, PPO-B, HRP-Ab) for the detection of cholera antitoxin. (A) HRP-immunosensor, (B) GOX-B-immunosensor, (C) PPO-B-immunosensor. Mred/Mox = hydroquinone/quinone Gox = biotinylated glucose oxidase PPO — biotinylated polyphenol oxidase HRP-Ab = peroxidase-labeled IgG anti-rabbit antibody.

Applications. The most common application of electrofunctional polymers is in the development of new materials in the (bio)sensors field [78]. For instance, a pyrrole-modified biotin allows successive attachment of avidin and biotin-labeled glucose oxidase, resulting in an efficient glucose biosensor [77,79]. Oureghi et al. [80] developed an impedimetric immunosensor for... 

Immunochemical analysis is a fast developing field with numerous possibilities for further improvement. Much effort is being put into the development of continuous measurements, such as flow-injection immunoanalysis (FI I A) and immunosensors. A quasicontinuous FI IA of pesticides was developed by Kramer and Schmid ) on the basis of a competitive lA. Here, the Abs are immobilized on a membrane. The reaction takes place in the membrane reactor, the central part of the flow injection system. All reagents are sequentially added to the reactor and the product is assayed with the aid of a fiow fiuorimeter. The measuring range of the fiow injection analysis almost equals that of the EIA. Wittmann and Schmid used an Ab column reactor filled with polystyrene or glass beads with the Ab immobilized via the avidin/biotin system. This system showed a stable Ab activity for a minimum of 500... 

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