Enzyme-linked immunosorbent assays direct competition

Kleivdal, H., Kristiansen, S.I., NUsen, M.V., Gokspyr, A., Briggs, L., Holland, F.T., and McNabb, R, Determination of domoic acid toxins in shellfish by Biosense ASP ELISA—a direct competitive enzyme-linked immunosorbent assay collaborative study. J. AOAC Int., 90, 1000, 2007. 

Hesp, B.R. et ah. Detection of domoic acid in rat serum and brain by direct competitive enzyme-linked immunosorbent assay (cELISA), Anal. Bioanal. Chem., 383, 783, 2005. 

Dixon-Holland, D. E. Katz, S. E. Direct competitive enzyme-linked immunosorbent assay for sulfamethazine residues in milk. J. Assoc. Off. Anal. Chem., 72 447-50. 1989. 

At their most elaborate, epitope mapping techniques can provide detailed information on the amino acid residues in a protein antigen, which are in direct contact with the antibody binding site. X-ray crystallography of antibody-antigen complexes can identify contact residues directly and unequivocally, though not surprisingly in view of the effort required, this method is not in routine use. At the other extreme, demonstration by competition enzyme-linked immunosorbent assay (ELISA) methods that two antibodies bind to different sites on... 

Competitive enzyme-linked immunosorbent assays (ELISA) Two types of ELISA have been used for the analysis of mycotoxins and both types are heterogenous competitive assays. One type, i.e. direct ELISA, involves the use of a mycotoxin-enzyme conjugate and the other system, i.e. indirect ELISA, involves the use of a protein-mycotoxin conjugate and a secondary antibody to which an enzyme has been conjugated. Although horseradish peroxidase (HRP) is most commonly used as the enzyme for conjugation, other enzymes such as alkaline phosphatase and beta-galactosidase, also have been used (5, 9, 13). 

The most widely used immunological approaches are those based on enzyme-conjugated antibodies, offered as kits that allow immunochemical detection of allergens and toxic components in foods. These products are popular in that they offer ease of automation, the possibility of analyzing a large number of samples in a single approach, besides relying on safe chemicals and simple protocols. Either direct or competitive enzyme-linked immunosorbent assay... 

In a direct immunoassay the immobilized antibody binds to the corresponding antigen. The competitive immunoassay relies upon the competition of the analyte with a labelled analyte for antibody binding. These formats are widely used for high throughput affinity arrays. A sandwich immunoassay is based on the trapping or capture of the analyte by another antibody. In ELISA (enzyme linked immunosorbent assays) the second antibody is conjugated with an enzyme. The bound enzyme labelled antibody is detected by its ability to break down its substrate to a colored product. 

Enzyme-linked immunosorbent assay (ELISA) is based on the specific reaction between an antibody and an antigen. One of the reagents in the reaction is labeled with an enzyme that generates a colorimetric product that can be measured with a spectrophotometric device. The color intensity correlates with the concentration of specific antibody and the respective antigen. The reaction can be formatted in various ways in a multiwell plate (microtiter plate) with the common formats being the sandwich assay, the competitive assay, and the direct assay. (See Figure 11.1.)... 

Enzyme-Linked Immunosorbent Assays (ELISA). Three methods are commonly used direct competition, double antibody sandwish and antibody inhibition. 

Immunoassay is well acknowledged since the introduction of ELISA (enzyme-linked immunosorbent assay) which was developed over 20 years ago. ELISA can be classified into three formats direct, sandwich, and competitive [2]. Direct ELISA is a simple process that the excessive antibody is reacted with antigen. After incubating, a washing step is conducted to eliminate the unbound antibody. The sensitivity is proportional to the amount of the present antibody in the solution. In a sandwich immunoassay, the antigen (e.g., sample) is between the primary antibody and the second antibody (detection antibody). Generally, the primary antibody is immobilized on the solid surface and the second antibody is labeled with certain enzyme or fluorophore. The amount of sample is proportional to the amount of the labeled second antibody which is measured by the different detection strategies. In a competitive immunoassay, the... 

Immunoassays vary by the different labels they use. The most common labels include chromophores, fluorophores, radioisotopes and enzymes. Of those labels, enzyme immunoassay or enzyme-linked immunosorbent assay (ELISA) is the most popular technique. ELISA has as an advantage the amplification of the analytical signal and/or increase of the sensitivity of the immunoassay. There are four types of ELISA direct ELISA, indirect ELISA, sandwich ELISA and competitive ELISA. 

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