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Pros and cons of immunological methods.
M. TILLEY (1), C. Don (2), K. Konitzer (3), P. Koehler (4). (1) USDA-ARS, Manhattan, KS, U.S.A.; (2) CDC Foodphysica, Driel, Netherlands; (3) Deutsche Forschungsanstalt für Lebensmittelchemie, Freising, Germany; (4) Deutsche Forschungsanstalt fuer Lebensmittelchemie, Freising, Germany

The International regulatory agencies Codex Alimentarius, European Commission Regulation and the U.S. Food and Drug Administration have set 20 mg/kg (ppm) as the maximum limit of gluten allowed in foods labeled as “gluten-free”. Immunological approaches appear to be, so far, the most suitable methods available in gluten detection. This is reflected by the R5 ELISA, which is listed as type I Codex method. ELISA has been the method of choice due to its low cost, ease of operation and specificity of antibodies. Considerable progress has been achieved in the development of monoclonal antibodies for commercial ELISA kits that are specific for common sequences in the different gluten bearing cereals or towards toxic sequences thus providing potential clinical significance. Although universally used there are several issues that must be considered. Analyte complexity is a major concern as gluten is not a single target molecule but an intricate network of numerous individual proteins that occur at different proportions within and among the different gluten-bearing cereals. Extraction is usually performed using aqueous alcohol, however food composition will impact analyte recovery so that reducing agents, gelatin and other agents are often required for complete gluten extraction. These issues have hampered the development of a universal reference standard. The Prolamin Working Group developed a well-characterized gliadin standard material, but total gluten is determined using a multiplication factor of 2 as the prolamin is considered to generally represent 50% of total gluten. The analysis of partially hydrolyzed gluten in fermented food is still an unsolved problem because it is not clear how to convert peptide into gluten concentrations. Peptic-typtic prolamin digests have been suggested as reference materials but there is no general acceptance of this type of calibrator.

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