Immunosensor construction

In the traditional format, screen-printed electrodes are used as solid phase to carry out the immunoassay and as signal transducers. In recent years, different kinds of beads (glass, graphite particles or magnetic particles) have been used as solid phase to perform the immunoassay. For this reason, it is possible to use a new approach in the immunosensor construction coupling beads to screen-printed electrodes. 

Based on preliminary photophysical end biological investigations a conclusion can be taken that the dendrimer membrane possesses promising properties for applying in biosensor and immunosensor constructions with fluorescent detection (Grabchev et al. 2007b) (Fig. 3). 

The EIS can provide useful information on the electron transfer resistance of the electrode surface during stepwise immunosensor construction process. The incorporation of nanomaterials and immobihzation of antibody onto the modified electrode surfaces were characterized by Faradaic EIS in the presence of the Fe(CN)6] redox probe. According to the standard complex function representation, impedance (Z) can be presented as the sum as a real (Z ) and an imaginary part (Z") components. The curve of the EIS includes a semicircular part and a linear part. The semicircle... 

The cyt c immunosensor construction using CNT platform is similar to GNP platform but the sensing platform features a CNT-tadored PPy/SPE instead of GNP-electrodeposited PPy/SPE. The incorporation of CNT onto the PPy/SPE was performed by incubating 10 pL of CNT solution (0.5 mL of 1.5 wt% nafion—ethanol solution containing 1 mg of CNT) on PPy/SPE surface, and then the solvent ethanol was evaporated in air to form a CNT/PPy/SPE. The immobilization of anti-cyt c onto the CNT/PPy/SPE was carried out as described before similarly to SAM/GNP/ PPy/SPE platform. 

Particularly attractive for numerous bioanalytical applications are colloidal metal (e.g., gold) and semiconductor quantum dot nanoparticles. The conductivity and catalytic properties of such systems have been employed for developing electrochemical gas sensors, electrochemical sensors based on molecular- or polymer-functionalized nanoparticle sensing interfaces, and for the construction of different biosensors including enzyme-based electrodes, immunosensors, and DNA sensors. Advances in the application of molecular and biomolecular functionalized metal, semiconductor, and magnetic particles for electroanalytical and bio-electroanalytical applications have been reviewed by Katz et al. [142]. 

C. Steegborn and P. Skladal, Construction and characterization of the direct piezoelectric immunosensor for atrazine operating in solution. Biosens. Bioelectron. 12,19-27 (1997). 

Since immunosensors usually measure the signals resulting from the specific immu-noreactions between the analytes and the antibodies or antigens immobilized, it is clear that the immobilization procedures of the antibodies (antigens) on the surfaces of base transducers should play an important role in the construction of immunosensors. Numerous immobilization procedures have been employed for diverse immunosensors, such as electrostatic adsorption, entrapment, cross-linking, and covalent bonding procedures. They may be appropriately divided into two kinds of non-covalent interaction-based and covalent interaction-based immobilization procedures. 

R.E. Ionescu, C. Gondran, L.A. Gheber, S. Cosnier, and R.S. Marks, Construction of amperometric immunosensors based on the electrogeneration of a permeable biotinylated polypyrrole film. Anal. Chem. 76, 6808-6813 (2004). 

Starodub et al. [98] studied different constructions and biomedical applications of immunosensors based on fiberoptic and enhanced CL. They discussed three different approaches of immobilization of one of the immunocomponents on the fiberoptic surface. Good results could be achieved by the use of a special membrane closely connected to the fiberoptic, with sensitivities compared to that obtained by the ELISA method but with a faster rate of analysis. The sensor was... 

Non-labelled immunosensors rely on various principles (Fig. 3.27.A). Either the antibody or the antigen is immobilized on the solid matrix to form a sensing device. The solid matrix should be sensitive enough at the surface to detect immunocomplex formation. Electrode, membrane, piezoelectric and optically active surfaces may in principle be used to construct non-labelled immunosensors. The antigen or antibody to be determined is dissolved in a solution and reacted with the complementary matrix-bound antibody or antigen to form an immunocomplex that alters the physical e.g. the electrode potential or intrinsic piezofrequency) or optical properties of the... 

Construction of amperometric immunosensors for the analysis of cholera antitoxin and comparison of the performances between three different enzyme markers... 

Two immunosensors developed by O Regan et al. [89,90] have demonstrated their usefulness for the early assessment of acute myocardial infarction (AMI). Human heart fatty-acid binding protein (H-FABP) is a biochemical marker for the early assessment of AMI. The authors constructed an amperometric immunosensor for the rapid detection of H-FABP in whole blood. The sensor is based on a one-step, direct sandwich assay in which the analyte and an alkaline phosphatase (AP) labelled antibody are simultaneously added to the immobilized primary antibody, using two distinct monoclonal mouse anti-human H-FABP antibodies. The substrate p-amino-phenyl phosphate is converted to p-aminophenol by AP, and the current generated by its subsequent oxidation at +300 mV vs. Ag/AgCl is measured. The total assay time is 50 min, and the standard curve was linear between 4 and 250 ng ml . The intra- and inter-assay coefficients of variation were below 9%. No cross-reactivity of the antibodies was found with other early cardiac markers, and endogenous substances in whole blood did not have an... 

Number of antibodies that can be produced is practically unlimited, hence wide development of immunoassays not only for determination of macromolecules, which induce immunoreactions, but also for small molecule analytes inducing such reaction after binding into suitable conjugates. Besides immunoassays carried out in various formats and immunoaffinity liquid chromatography, since many years very broad studies are carried out on design of integrated immunosensors.127129 In principle their construction should simplify the immunochemical determination by elimination various steps necessary in conventional immunoassays based on the use of labels to monitor the immunochemical reaction. In immunochemical process, the... 

Lemmer K, Donoso Mantke O, Bae HG, Groen J, Drosten C, Niedrig M (2004) External quality control assessment in PCR diagnostics of dengue virus infections. J Clin Virol 30(4) 291-296 Leshem B, Sarfati G, Novoa A, Breslav I, Marks RS (2004) Photochemical attachment of biomolecules onto fibre-optics for construction of a chemiluminescent immunosensor. Luminescence 19(2) 69-77... 

Horseradish peroxidase (HRP) is an important peroxidase that contains heme, which is the protein active site with the resting state of the heme iron, Fe(III), as prosthetic group. It can catalyze the dependent one-electron oxidation of a great variety of substrates, and has been commonly employed to construct biosensors (Hai-Li Zhang et al. 2008). Peroxidase is the most frequently used enzyme for the construction of immunosensors. Successful immobilization of horseradish peroxidase on modified magnetic particles and their employment in the amperometric biosensors are mentioned in (Hai-Li Zhang et al. 2008 Yu et al. 2006). 

Fig. 9. Reflectometric immunosensor. (a) Construction scheme (b) dependence of the measuring signal on layer thickness upon reaction between adsorbed immunoglobulin G (IgG) and anti-IgG. (Gelatin is added to suppress nonspecific adsorption on the free silicon surface.) (Redrawn from Welin et al., 1984).

Since in most cases only one enantiomer possesses a desired pharmacological activity, it is necessary to construct enantioselective sensors to improve the quality of analysis due to the high uncertainty obtained in chiral separation by chromatographic techniques.315 For this purpose, enantioselective amperometric biosensors and potentiometric, enantioselective membrane electrodes have been proposed.264 The selection of one sensor from among the electrochemical sensor categories for clinical analysis depends on the complexity of the matrix because the complexity of different biological fluids is not the same. For example, for the determination of T3 and T4 thyroid hormones an amperometric biosensor and two immunosensors have been proposed. The immu-nosensors are more suitable (uncertainty has the minimum value) for direct determination of T3 and T4 thyroid hormones in thyroid than are amperometric biosensors. For the analysis of the same hormones in pharmaceutical products, the uncertainty values are comparable. 

---