Application Guide - QIAxcel Advanced

116 QIAxcel Advanced Application Guide 10/2016 Identifying allergenic nut species using the QIAxcel® system Malcolm Burns Molecular Biology, LGC, Queens Road, Teddington, Middlesex TW11 0LY, UK Introduction The global food industry faces numerous challenges in terms of providing confidence in the authenticity of foods. There is a commercial interest in ensuring that foods are correctly labeled for the presence and levels of specific ingredients, including the occurrence of GMOs and incidence of allergens, and in preventing the fraudulent replacement of expensive food ingredients with inferior ones. For the benefit of consumers who are allergic to certain food ingredients, current EU regulations require labeling disclosure of the presence of any of 14 major allergens if used as ingredients in prepacked foods (Directive 2003/89/EC, as amended). PCR assays have been developed to detect and identify DNA originating from allergenic nut materials in processed foods. They consisted of a suite of singleplex and multiplex assays to detect DNA from almond, hazelnut, macadamia nut, and peanut. The assays have been validated by LGC (UK) and Premier Analytical Services (UK) with regards to sensitivity and specificity. This study demonstrates the successful use of the QIAxcel system for the analysis of PCR products as part of an allergen detection workflow. The QIAxcel system provided rapid, reliable, and inexpensive identification of four nut species. Materials and Methods Samples from a panel of 17 food products (Table 1) were analyzed using singleplex and multiplex PCR assays developed for the nut species almond, hazelnut, macadamia nut, and peanut. The PCR products were analyzed using the QIAxcel system in combination with the QIAxcel DNA High Resolution Kit. End-point PCR was conducted for each nut species using 200 µM primers and 50 ng template DNA in a final volume of 25 µl. The PCR products were in the size range 75 to 134 bp. Short fragment sizes were chosen to ensure that the targets remained intact during food processing. The DNA concentration of the samples tested was in the range 2–5 ng/µl. Samples were analyzed using a QIAxcel DNA High Resolution Cartridge. The method designated OM500 was chosen as the optimal one for the nut species under analysis. It has the following parameters: QX Alignment Marker injection at 4 kV and 20 s, sample injection at 5 kV for 10 s and separation at 5 kV for 500 s. The sample injection time was increased to 40 s to ensure identification of all of the amplicons present in the sample. The analysis was performed with a suspend integration time of 2.53 min. QX Alignment Marker 15 bp/400 bp and QX DNA Size Marker 25–500 bp were run simultaneously. Two fragments from the QX DNA Size Marker were excluded from the analysis since they were longer than the 400 bp fragment in the QX Alignment Marker.

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