(585v) In Vitro Cell-Based Assays for Assessing the Impact of Thermal Processing On the Allergenic Potential of Milk Allergens

A better understanding of how processing may affect the allergenicity of proteins will allow a more accurate allergen risk assessment and achieve a greater success in food allergy management. To determine whether food processing can affect the allergenic potential of proteins, immunochemical methods such as ELISA are commonly used, however, these methods are not able to determine changes in the proteins’ ability to elicit allergic reactions. Although food challenge studies are the gold standard for determining the allergenicity of proteins, such studies are seldom performed due to ethical reasons and the difficulty in recruiting a sufficient number of study participants. In recent years, a number of mediator release assays have been developed to measure the allergenic potential of food proteins in vitro. How these cell-based assays can be used to assess the impact of thermal or other processing conditions on the allergenic potential of proteins remains to be determined. This study evaluated the use of sensitized rat basophilic leukemia (RBL) cells for assessing the impact of boiling on the allergenic potential of milk allergens by measuring changes in the proteins’ ability to induce cell degranulation and the release of β-hexosaminidase. RBL-2H3 cells sensitized with rat IgEs that were specifically raised against NIST non-fat milk standard reference material, β-lactoglobulin (BLG), α-lactalbumin (ALA) and α-casein (αCAS) were challenged with various concentrations of each allergen in its native and processed (boiled in PBS for 10 min) forms. Cell degranulation caused by the allergen was monitored by measuring the release of β-hexosaminidase. All allergens (NIST milk, BLG, ALA, and αCAS) were able to induce the release of β-hexosaminidase release, ranging from 7.90% to 39.02%. In general, a decrease in % release of β-hexosaminidase was observed when the cells were challenged with a decreasing concentration of the allergens. Boiling of ALA caused an increase in the % release of β-hexosaminidase from sensitized cells across antigen concentrations from 10^-9 mg/mL to 1 mg/mL. In conclusion, this study demonstrated that the in vitro mediator release assay method may serve as a useful model to evaluate the impact of thermal processing on the allergenic potential of milk allergens.