Abstract
BackgroundOil body-associated allergens such as oleosins have been reported for important allergenic foods such as peanut, sesame and hazelnut. Here we investigate whether oil body associated proteins (OAPs) are linked with specific clinical phenotypes and whether they are represented in skin prick test (SPT) reagents.MethodsA hazelnut OAP fraction was characterized by mass-spectrometry (MS) to identify its major constituents. Polyclonal rabbit antibodies were generated against hazelnut OAPs. The presence of OAPs in commercially available hazelnut SPTs was studied by immunoblot and spiking experiments. OAP-specific IgE antibodies were measured in sera from patients with a convincing history of hazelnut allergy by RAST (n = 91), immunoblot (n = 22) and basophil histamine release (BHR; n = 14).ResultsHazelnut OAPs were analysed by MS and found to be dominated by oleosins at ~14 and ~17 kDa, and a 27 kDa band containing oleosin dimers and unidentified protein. In 36/91 sera specific IgE against hazelnut OAPs was detected, and confirmed to be biologically active by BHR (n = 14). The majority (21/22) recognized the oleosin bands at 17 kDa on immunoblot, of which 11 exclusively. These OAP-specific IgE responses dominated by oleosin were associated with systemic reactions to hazelnut (OR 4.24; p = 0.015) and negative SPT (χ2 6.3, p = 0.012). Immunoblot analysis using OAP-specific rabbit antiserum demonstrated that commercial SPT reagents are virtually devoid of OAPs, sometimes (3/9) resulting in false-negative SPT. Spiking of SPT reagents with OAP restored serum IgE binding of these false-negative patients on immunoblot at mainly 17 kDa.ConclusionHazelnut allergens found in oil bodies dominated by oleosin are associated with more severe systemic reactions and negative SPT. Defatted diagnostic extracts are virtually devoid of these allergens, resulting in poor sensitivity for detection of IgE antibodies against these clinically relevant molecules.
Highlights
Oil body-associated allergens such as oleosins have been reported for important allergenic foods such as peanut, sesame and hazelnut
Oleosins are embedded in the oil body with a highly conserved hydrophobic central domain [24], whereas the hydrophilic alpha-helical N-terminus and amphipathic alpha-helical C-terminus, which is conserved among many oleosins [20,21,22], are in contact with the aqueous cytoplasm [25].Oleosins have been identified as allergens in peanut, sesame seed and hazelnut [16,26,27]
The aim of this study was to test commercially available skin prick test (SPT) reagents with respect to the presence of Oil body associated protein (OAP), and to use purified OAPs from hazelnut to evaluate the clinical importance of OAP-specific IgE antibodies using sera from well-characterized patients with a convincing history and/or positive Double-blind placebo-controlled food challenge (DBPCFC) for hazelnut
Summary
Oil body-associated allergens such as oleosins have been reported for important allergenic foods such as peanut, sesame and hazelnut. Primary sensitization to the major birch pollen allergen Bet v 1 results in cross-reactivity to globulin), the hazelnut storage proteins Cor a 11, Cor a 14 and Cor a 9 respectively [10,11,12] The latter two have recently been demonstrated to be associated with more severe objective symptoms in hazelnut allergy[11,13]. The low levels of LTP can be explained by the extraction protocols used for peanut or hazelnut, which are commonly carried out at neutral pH where LTPs are optimally soluble at acidic pH [18,19] Another characteristic of extraction procedures used for legumes, seeds and nuts is that defatting steps are included. Oleosins are embedded in the oil body with a highly conserved hydrophobic central domain [24], whereas the hydrophilic alpha-helical N-terminus and amphipathic alpha-helical C-terminus, which is conserved among many oleosins [20,21,22], are in contact with the aqueous cytoplasm [25].Oleosins have been identified as allergens in peanut, sesame seed and hazelnut [16,26,27]
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