Quantitative Sandwich ELISA for the Determination of Fish in Foods

Christiane K. Fæste, Christin Plassen, Lise Holden, Marianne T. Werner

National Veterinary Institute, PO Box 8156 dep., N-0033 Oslo, Norway

Fish is among the most common causes of IgE-mediated food allergy in Europe. Small amounts of fish protein can cause life-threatening reactions as has been proven by food provocation studies. The major allergen in fish is parvalbumin, an ubiquitous calcium-binding muscle protein of about 12 kDa, which is highly conserved in different fishes and probably is responsible for the extensive clinical cross-reactivity observed. A number of parvalbumins (e.g. from cod and salmon) have been characterised on molecular level. They contain sequential epitopes which may explain their considerable stability to food processing methods like heating, pH-shifts or denaturing.

Detection of fish in food has previously been based on the use of patient serum. We have developed a novel sandwich enzyme-linked immunosorbent assay (ELISA) for the quantitation of fish in processed foods, using a polyclonal anti-cod parvalbumin (Gad m 1, Atlantic cod) antibody for capture and a biotinylated conjugate of the same antibody for detection. The antibody was specific for fish. Cross-reactions to other common foods were not observed. The assay had a detection limit of 0.28 mg/kg in matrices like bread, wine, soy sauce, white sauce, and soup. Recoveries were determined at 1, 10, and 100 mg/kg and ranged from 58-127 %, while the intra-assay and inter-assay coefficients of variation were < 10% and < 19%, respectively.