Multiplex ligation-dependent probe amplification (MLPA) for simultaneous detection of DNA from sunflower, poppy, flaxseed, sesame and soy allergenic ingredients in commercial food products

Abstract

This work describes a multiplex ligation-dependent probe amplification (MLPA) technique for the simultaneous detection of five food allergens: sunflower, poppy, flaxseed, sesame and soy. Species-specific MLPA half-probes were designed to detect the DNA from the targeted species. Ligated probes were amplified by polymerase chain reaction (PCR) and amplicons were detected using capillary electrophoresis. The specificity of the MLPA system was assessed against DNA from more than 50 plant and animal species. The limit of detection (LOD) of the assay was determined to be 10 mg/kg in a model cookie experimentally spiked with different concentrations of the target species. The applicability of the MLPA was demonstrated through the analysis of 56 different commercial products (breads, pastries, cereals, chocolates, drinks, etc.), evidencing the presence of one or more undeclared allergenic ingredients in some of the tested samples. Real-time PCR assays were also performed to confirm and complement MLPA results. The MLPA technique has proved as a qualified screening tool for determining the presence of low amounts of sunflower, poppy, flaxseed, sesame and soy in processed foods.