Detection of fish protein in processed food Haiyan Min, Kamil Gajewski and Yun-Hwa Peggy Hsieh Abstract The Food Allergen Labeling and Consumer Protection Act (FALCPA) of 2004, requires mandatory labeling of the eight major food allergens (fish, shellfish, milk, egg, soy, wheat, peanuts and tree nuts) for pre-packaged foods. To enforce the labeling law, method development for detecting undeclared allergens in food is important because reliable analytical methods for detection of the ingredients derived from these eight foods can significantly reduce the risk of allergic reactions. Currently no methods are available for the detection of the presence of fish proteins in processed food. In the present study, an indirect non-competitive enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies (Mabs) was developed for this purpose. The soluble protein from 45 raw and cooked fish species were separated by sodium dodecyl sulfat polyarylamide gel eletrophoresis (SDS-PAGE), and the antigenic components were examined by Western blot using two MAbs: 2G8 and 3E4, which were previously developed in our laboratory against heated crude catfish proteins. Indirect noncompetitive ELISA was employed to discriminate between cooked fish samples and nine meat samples (pork, beef, lamb, chicken, turkey, rabbit, deer, elk, and horse). The lab adulterated samples of imitation crab sticks (a surimi product made from Alaska Pollock) in pork and in chicken were also analyzed by these two Mabs. Mabs 2G8 and 3E4 reacted to all the 45 fish and shellfish species tested, including majority of the top ten most consumed seafood in the US (shimp, tuna, salmon, pollock, tilapia, catfish, and cod). Western blot revealed that Mabs 3E4 and 2G8 recognized several thermal-stable antigenic proteins (12, 18, and 25 kDa) in the fish extracts. The ELISA results confirmed that the Mabs only reacted with fish and shellfish samples, but not the meat samples tested. These Mabs (2G8 and 3E4) can also distinguish surimi products from chicken and pork samples, with the detection limit of 1% and 2% (w/w), respectively. Surimi products may be a hidden ingredient in some processed foods, which may induce allergic reactions in sensitive individuals. The immunoassay developed is useful for the detection of low levels of undeclared fish proteins in raw and heat-processed food products.
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