D2.08 - Extraction Efficiency Studies and Quantification of Major Peanut Allergens in Peanut Challenge Meals and Peanut Flour

Extraction efficiency of Ara h 1, Ara h 2, Ara h 3 and Ara h 6 from different matrices will have a direct effect on the measured allergen content. This study looks to identify the optimum extraction buffer and procedure for component peanut allergens Ara h 1, Ara h 2, Ara h 3 and Ara h 6 from 3 different matrices: 2 peanut challenge meal (PCM) formulations (each at 3 different strengths) and peanut flour.

Three extraction buffer compositions were studied per peanut component allergen in peanut flour and PCM 6months+/4years+ (low and high strength) and 2 for the 0% w/w PCM formulations. Allergen levels were quantified, using allergen-specific ELISA. For the PCMs, matrix-recovery of component allergens was calculated using average specific allergen content determined in peanut flour, adjusted to %w/w peanut flour.

PCM matrix-recovery of Ara h 2, Ara h 3 and Ara h 6 from PCM 4y+ (low and high strength) were within 82% to 101%. Ara h 1 recovery was lower at 1% to 32%. For PCM 6m+ (low and high strength) recoveries were within 91% to 138%. Allergen was below method limit of detection in the 0% w/w PCM samples. Extraction replicates had highly agreeable allergen content, with an average %CV of 5-13%. 

Extraction buffers have been identified for Ara h 1, Ara h 2, Ara h 3 and Ara h 6 across multiple matrices. The data suggests the extraction and analytical methodology are robust and repeatable and the challenge meals are uniform in allergen content. This is important for quality, safety and efficacy controls in standardising materials for allergy diagnosis.