Development and validation of reference material for the detection of cashew nut ingredients by real-time quantitative PCR

To solve the problem of the lack of reference material (RM) for determination of allergenic ingredients in food, a RM of cashew nut powder was developed in the study. Cashew nut powder was prepared from cashew nut kernel by selecting, cleaning, crushing, n-hexane degreasing and sieving treatment. The reliability and traceability of RM was verified using real-time quantitative polymerase chain reaction (qPCR) and phylogenetic tree analysis. The cashew nut powder RM showed good homogeneity, and good stability under long-term storage at 4 ℃ and short-term simulated transportation from –20 to 45 ℃. The RM was determined jointly by 8 collaborative laboratories, and the characteristic C_T value was 24.732, and the extended uncertainty was 1.052% (k = 2). The RM was applied to verify the amplification efficiency and the limit of detection for qPCR assay, and showed good applicability. The RM could be used for method validation and quality control, for the determination of allergenic ingredients in food mixed with trace amounts of cashew nut.