Development of a droplet digital Polymerase Chain Reaction assay for peanut detection of food allergens

Abstract

Food allergies are a risk for patients, especially peanut-allergy, even at low concentration
it is still effective. As a consequence, a regulation based on the precautionary principle
requires the labelling to declared allergenic ingredients. This enforces an importance
to develop and validate the appropriate methods in accurately detection of peanut DNA.
Therefore, the aim of the present work was to validate the method in order to detect and
quantify peanut DNA by using droplet digital PCR (ddPCR). Peanut DNA was extracted with
DNeasy mericon Food Kit (Qiagen) and was used as the template to optimise method. The
assay was designed to target Arachis hypogaea allergen II gene based on hydrolysis probe.
ddPCR was used to detect and quantify peanut DNA without the requirement of calibration
curve. The limit of detection (LOD) was 0.015 ng/μL or 3.04±0.82 copies/μL and the limit of
quantification (LOQ) was 0.03 ng/μL or 9.21±1.11 copies/μL. It can be said that the ddPCR
assay was sensitive, reliable and could be used as a quantitative tool for detection of
allergenic-peanut.