Direct competitive ELISA

Another commonly used ELISA format is the immobilized antibody assay or direct competitive assay (Eigure 3). The primary anti-analyte antibody is immobilized on the solid phase and the analyte competes with a known amount of enzyme-labeled hapten for binding sites on the immobilized antibody. Eirst, the anti-analyte antibody is adsorbed on the microtiter plate wells. In the competition step, the analyte and enzyme-labeled hapten are added to microtiter plate wells and unbound materials are subsequently washed out. The enzyme substrate is then added for color production. Similarly to indirect competitive immunoassay, absorption is inversely proportional to the concentration of analyte. The direct competitive ELISA format is commonly used in commercial immunoassay test kits. 

Regarding commercially available immunochemical kits, we could mention the Charm ROSA Enrofloxacin Test that detects ciprofloxacin and enrofloxacin equally (see Table 4) and the 5101ERFXlp test. This last one is a direct competitive ELISA, which uses MAbs and has a LOD of 3 ng g1 in tissues. Some other companies do have antibodies available as reagents for different applications such as Biodesign International and QED Bioscience Inc. 

This method is further divided into two major types. One type is competitive ELISA, which can be used for the analysis of both hapten and macromolecule the other is noncompetitive sandwich-type ELISA, which is only used for divalent and multivalent antigens. Two major types, i.e., direct competitive ELISA (dC-ELISA) and indirect competitive ELISA (inC-ELISA), are used most commonly in food analysis. 

In the direct competitive ELISA, the analyte specific antibodies are first coated on a solid phase. The sample or standard solution of analyte is generally incubated simultaneously with the analyte enzyme conjugate or incubated separately in two steps. The amount of enzyme bound to the plate is then determined by incubation with a chromogenic substrate solution. The resulting color/fluorescence, which is inversely proportional to the analyte concentration present in the sample, is then measured instrumentally or by visual comparison with the standards. 

The ETA for mycotoxins has been used since the late 1980s. It is based on the principle of the direct competitive ELISA. Anti-mycotoxin antibody is coated on the surface of a membrane. 

Direct Competitive ELISA for Antigen Detection and Quantification... 

ELISA methods for aflatoxins assay have been available for more than a decade. The technology is based on the ability of a specific antibody to distinguish the three-dimensional structure of specific aflatoxins. The direct competitive ELISA is commonly used in aflatoxin analysis [74],... 

A typical principle of direct competitive ELISA is shown in Figure 11.3. After an aflatoxin is extracted from a ground sample with solvent, a portion of the sample... 

Ridascreen -Folsaure, Direct competitive ELISA enzymoinmunoassay for fohc acid (Ref. 16311), r-biopharm, ATOM, Digen Ltd., U.K., 1998. 

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