Validating allergen coding genes (Cor a 1, Cor a 8, Cor a 14) as target sequences for hazelnut detection via real-time PCR

Hazelnuts have been shown to contain different allergenic proteins. Amongst these, major allergens Cor a 1 and Cor a 8 and the 2S albumin Cor a 14 were selected as targets to comparatively validate three Real- Time PCR protocols. We investigated both on the choice of the amplification target, and on the matrix effect on different sample foods. Applying statistics on the validation parameters obtained from the three protocols, we showed a significant difference in Ct values. This could turn critical when a high sensitivity method is required for the detection of hazelnut traces, confirming how fundamental the choice of the template during primer design phase is. Concluding, statistical approach represents a useful tool for the identification of the best performing primer pairs in Real-Time PCR. Cor a 8 gene permitted the iden- tification of hazelnut based ingredients in complex foods, providing a significantly higher sensitivity in the PCR amplification, when compared to Cor a 1 and Cor a 14.