Selective cloning of peanut allergens, including profilin and 2S albumins,by phage display technology

Background: Peanut kernels contain many allergens able to elicit IgE-mediat ed type 1 allergic reactions in sensitized individuals. Sera from sensitize d patients recognize variable patterns of IgE-binding proteins. The identif ication of the IgE-binding proteins of peanut extract would faciliate impro vement of diagnostic and immunotherapeutic approaches as well as developmen t of sensitive test systems for the detection of hidden peanut allergens pr esent as additives in various industrial food products and the investigatio n of their stability during processing of food products. Methods: We applie d the pJuFo cloning system based on the phage surface display of functional cDNA expression products to clone cDNAs encoding peanut allergens. Sera (n = 40) of peanut-allergic individuals were selected according to case histo ry, radioallergosorbent test and immunoblot analysis to demonstrate IgE bin ding towards the newly identified recombinant allergens. Results: In additi on to the known allergens Ara h 1 and Ara h 2 we were able to identify four allergens with estimated molecular weights of 36, 16, 14.5 and 14 kDa. Thr ee of them formally termed Ara h 4, Ara h6 and Ara h 7 show significant seq uence similarities to the family of seed storage proteins and the fourth (A ra h 5) corresponds to the well-known plant allergen profilin. Immunoblotti ng of the six expressed recombinant allergens with 40 patients sera shows 1 4 individual recognition patterns and the following frequency of specific I gE binding: Ara h 1 was recognized by 65%, Ara h 2 by 85%, Ara h 4 by 53%, Ara h 5 by 13%, Ara h 6 by 38% and Ara h 7 by 43% of the selected sera. Con clusions: All of the selected peanut-positive sera can detect at least one of the six identified recombinant allergens which can be used to establish individual patients' reactivity profiles. A comparison of these profiles wi th the clinical data will possibly allow a further insight into the relatio nship between clinical severity of the symptoms and specific IgE levels tow ards the six peanut allergens.