Chestnut allergen detection in complex food products: Development and validation of a real-time PCR method

Chestnut is gaining importance as food allergen, but a specific detection method has not been performed so far. For the first time, the development of a real-time PCR assay for chestnut allergen detection in processed foods is described. Initially, three sets of novel primers specific for allergen-encoding genes (Cas s 5, Cas s 9 and Cas s TLP) were designed and evaluated for chestnut detection. Primers for Cas s 9 and a TaqMan probe were finally selected for a sensitive and specific detection of chestnut in binary mixtures with a practical limit of detection (LOD) of 100 ppm. Effect of processing has been evaluated and sensitivity slightly reduced after boiling treatment. Amplification of Cas s 9 was importantly affected when heat and pressure were applied (autoclaving). Applicability of the method was tested by analysis of commercial food products. Consequently, the results support the development and validation of a real-time PCR method for accurate chestnut allergen detection and labelling.