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Validation of enzyme linked immunoabsorbant assays (ELISA) for the determination of latex allergens in food contact materials and associated foods
Project Code: A03056
Leatherhead Food International
Topping, J ; Haines, J
Natural rubber latex (NRL) is the milky sap derived from rubber trees. It is used to manufacture many goods a few of which are used in the food chain. Some individuals are allergic to the naturally occurring proteins present in the sap. Although exposure to latex allergens more often occurs in a healthcare setting, there have been occasional case reports in the literature of individuals with latex allergy experiencing allergic reactions after coming into contact with food wrappers sealed with latex based adhesive or after eating food prepared by food handlers wearing latex gloves. Therefore, the Food Standards Agency commissioned work to determine the extent of usage of natural rubber latex containing food contact materials and to develop quantitative methods to measure latex allergens in foods.
Previous work (FSA Project A03043) established that latex allergens could be detected in extracts from some food contact materials. Commercial ELISAs for four clinically relevant latex allergens: Hev b1, Hev b3, Hev b5 and Hev b6.02 were set up and a modified protocol developed which improved the detection of low levels of allergen. Recovery of allergens from adhesive spiked food samples was variable: good recovery was reported for Hev b5 and Hev b6.02 from most matrices but the recovery of Hev b3 and Hev b1 from spiked foods was low and variable. Preliminary experiments indicated that transfer of allergen to food from food contact materials was possible.
In this current report, several formulations of NRL-based cold seal adhesive and five batches of bakery release film were tested for their latex allergen content. These contact materials were chosen for study in this project over transfer from latex gloves since contact with gloves would be transient and it was felt that direct food contact should be given priority. The profile of allergens was different between the two types of contact material tested and there was a wide variation in the concentration of allergen between different batches of the same material.
Our previous report noted that some allergens were difficult to recover from some food matrices, so in this project various approaches including sonication, use of organic polar solvents and detergents and lipase digestion, were tested to try and improve the recovery of latex allergens from spiked food samples. Despite investigating a number of different methods, it has not been possible to improve the recovery of either Hev b1 or Hev b3 from adhesive spiked foods. However, latex allergen ELISAs for Hev b5 and Hev b6.02 were successfully validated in-house and then tested at a second laboratory with comparable results. The reported recoveries for Hev b5 from two food matrix categories, chocolate confectionery and frozen confectionery were 68 ± 6% and 83 ± 3% respectively. The recoveries for Hev b6.02 were 92 ± 9% and 96 ± 8% respectively for the same two food matrix categories. An unexpected finding in this study was the cross-reactivity of several flour samples in all of the latex allergen ELISAs. Significant cross-reactivity was noted to gram flour (chickpea), wheat flour and rice flour. This affected the testing of food types such as biscuits and pastries. When applying the allergen ELISAs to foods therefore, the analysis should be limited to foods which do not contain these ingredients or are unlikely to have been crosscontaminated with these ingredients.
An inter-laboratory comparison of the method for measuring latex allergens Hev b5 and Hev b6.02 was undertaken; good agreement in results was obtained between the two laboratories. Whilst small scale and thus fairly superficial this did serve to give some confidence that the method can give comparable results when used in a different laboratory.
The validated method has been applied to wrappers and foods collected from manufacturing sites. Hev b5 was measured on 1 of the 3 wrappers tested and Hev b6.02 on all of the 3 wrappers tested. None of the foods, which were sampled from the areas most likely to be in contact with the cold seal adhesive, tested positive suggesting that the allergens had not transferred at levels above the detection limit of the method.
Using the methods established in this project, it should be possible to measure latex allergen concentrations in food contact materials and Hev b5 and Hev b6.02 allergens in some types of food samples. However, the clinical relevance of the concentrations of allergen measured and the current limits of detection need to be assessed in relation to clinical data determining the amount of allergen required to cause allergic reactions in susceptible individuals.
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