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Realised by ALMS™
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Abstract No.: MoP-014
Session: Food and Nutrition
Presentation date: Mon, Aug 28, 2006
Presentation time: 14:30 – 16:00

A Confirmation Method for Hidden Peanut Allergens in Food

Monique Bremer1, Dick Hooijerink1, Dion Luykx1, Jan Cordewener2, Twan America2, Rob Frankhuizen1

1 RIKILT-Institute of Food Safety, Wageningen, Netherlands
2 PRI - Plant Research International, Wageningen, Netherlands

Correspondence address: Monique Bremer, RIKILT-Institute of Food Safety, Biomolecular Detection, Bornsesteeg 45, Wageningen, 6708 pd Netherlands.

Keywords: Affinity, Chromatography; Chromatography, Liquid -, Nanoscale Capillary; MS/MS, Liquid Chromatography; Protein Identification.

Novel aspect: LC-MS/MS for detection of allergens in food.

 

Food allergy is caused by unwanted immune reactions in some individuals to certain proteins in food. The intake of even small amounts of these allergenic proteins can already induce adverse and sometimes fatal reactions. Therefore, hidden allergens in foods, for instance as a result of cross-contamination in food-producing facilities, pose a real threat to allergenic people. In order to protect the health and safety of allergic consumers labeling of the major food allergens in products has become compulsory from November 2005. To comply with regulations and to improve consumer safety, the need for methods that can detect trace amounts of allergenic proteins in foods is evident. So far, much attention has been paid to develop screening methods in particular immunoassays. The specificity of immunoassays relies on the specificity of the antibodies used. However, it is generally known that antibodies may react (to minor extent) with other proteins as well (so called cross-reactivity). As food products contain a large variety of proteins, false-positive test results can not be ruled out entirely when using immunoassays. Therefore, the development of confirmation methods that can identify the offending allergenic protein in food is needed.
In this study a confirmation method for the detection and identification of peanut allergens in food based on LC-MS/MS is developed. This method uses immunoaffinity chromatography as an enrichment and clean-up step prior to LC-MS/MS analysis. For the affinity purification monoclonal antibodies directed against peanut allergens are coupled to sepharose. The affinity purified peanut proteins are reduced, alkylated and trypsin digested. Subsequently, the peptide mixture is separated and analysed by nano RP-HPLC ESI-MS/MS. Database search identified a number of peanut allergens with high sequence coverage and peptides unique for peanut that were not identified in any other known protein sequence. To evaluate the utility of this method, peanut proteins were spiked into food matrices at different levels. Preliminary results show that peanut concentrations in the low range ppm range can be detected and identified.