Relevance of Ara h1, Ara h2 and Ara h3 in peanut‐allergic patients, as determined by immunoglobulin E Western blotting, basophil–histamine release and intracutaneous testing: Ara h2 is the most important peanut allergen

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Abstract
Background A number of allergenic proteins in peanut has been described and the relative importance of these allergens is yet to be determined. Objectives We have investigated the relevance of previously identified peanut allergens in well‐characterized peanut‐allergic patients by in vitro, ex vivo and in vivo assays. Methods Thirty‐two adult peanut‐allergic patients were included based on careful and standardized patient history and the presence of peanut‐specific IgE. The diagnosis peanut allergy was confirmed using double‐blind placebo‐controlled food challenges in 23 patients. Major peanut allergens Ara h1, Ara h2 and Ara h3 were purified from peanuts using ion‐exchange chromatography. IgE immunoblotting was performed and IgE‐cross‐linking capacity was examined by measuring histamine release (HR) after incubating patient basophils as well as passively sensitized basophils with several dilutions of the allergens. Intracutaneous tests (ICTs) using 10‐fold dilution steps of the purified allergens and crude peanut extract were performed. Results Ara h2 was recognized most frequently (26 out of 32) in all tests and induced both positive skin tests and basophil degranulation at low concentrations, whereas Ara h1 and Ara h3 were recognized less frequently and reacted only at 100‐fold higher concentrations as analysed with HR and intracutaneous testing (ICT). Next to the three tested allergens, proteins with molecular weights of somewhat smaller than 15 kDa were identified as a IgE‐binding proteins on immunoblot in the majority of the patients (20 out of 32). Conclusion We conclude that Ara h2 is, for our patient group, the most important peanut allergen, and that previously unidentified peanut proteins with molecular weights of somewhat smaller than 15 kDa may be important allergens as well. ICT in combination with basophil–HR and IgE immunoblotting provides insight in the patient specificity towards the individual peanut allergens.

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