Specific IgE to peanut 2S albumin Ara h 7 has a discriminative ability comparable to Ara h 2 and 6 (original) (raw)
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Clinical <html_ent glyph="@amp;" ascii="&"/> Experimental Allergy, 2004
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 wellcharacterized 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.
The Journal of Allergy and Clinical Immunology, 2003
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 wellcharacterized 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.
Clinical & Experimental Allergy, 2013
Background Several studies have analysed the diagnostic value of specific IgE (sIgE) for individual peanut allergens. However, little is known about the concordance between different techniques available in both children and adults. Objective To evaluate the value of individual peanut allergens by different techniques, i.e. multi-plexed microarray, single-plexed IgE assay, skin prick test (SPT) and immunoblot in both peanut allergic adults and children. Methods Sensitization patterns to peanut allergens Ara h 1, 2, 3, and 8 were evaluated using four different techniques: multi-plexed microarray immunoassay, single-plexed IgE assay, SPT and immunoblot. Twenty-two peanut allergic adults and 15 children scored on clinical severity according to double-blind, placebo-controlled food challenges and 27 atopic control patients were included. Results Comparable sensitivity values were found between all four techniques in adults, with the highest sensitivity for Ara h 2 (76.2-95.5%, compared to 100% with all techniques in children). The multi-plexed assay to Ara h 1 (93.3%) demonstrated a higher sensitivity compared with the other three techniques (P = 0.04) in children, but absolute values were perfectly correlated. There were no differences between adults and children. The area under the receiver operating characteristic curve (AUC) of sIgE to Ara h 1 was higher with the multi-plexed assay compared with the single-plexed assay (0.91 vs. 0.75). In adults, sIgE to Ara h 1, 2, and 3 was correlated with clinical severity. No such correlation was found in children. Conclusion and Clinical Relevance In conclusion, the single-and multi-plexed assay, SPT and immunoblot perform equally in both peanut allergic adults and children, with Ara h 2 being most often recognized with all techniques. Specific IgE to Ara h 1, 2, and 3 in adults was correlated with severity.
Journal of Clinical …, 1995
Peanut allergy is a significant health problem because of the frequency, the potential severity, and the chronicity of the allergic sensitivity. Serum IgE from patients with documented peanut hypersensitivity reactions and a peanut cDNA expression library were used to identify clones that encode peanut allergens. One of the major peanut allergens, Ara h I, was selected from these clones using Ara h I specific oligonucleotides and polymerase chain reaction technology. The Ara h I clone identified a 2.3-kb mRNA species on a Northern blot containing peanut poly (A)+ RNA. DNA sequence analysis of the cloned inserts revealed that the Ara h I allergen has significant homology with the vicilin seed storage protein family found in most higher plants. The isolation of the Ara h I clones allowed the synthesis of this protein in E. coli cells and subsequent recognition of this recombinant protein in immunoblot analysis using serum IgE from patients with peanut hypersensitivity. With the production of the recombinant peanut protein it will now be possible to address the pathophysiologic and immunologic mechanisms regarding peanut hypersensitivity reactions specifically and food hypersensitivity in general. (J. Clin.
Isoforms of the Major Peanut Allergen Ara h 2: IgE Binding in Children with Peanut Allergy
International Archives of Allergy and Immunology, 2004
Background: The major peanut allergen Ara h 2 consists of two isoforms, namely Ara h 2.0101 and Ara h 2.0201. The recently identified Ara h 2.0201 isoform contains an extra 12 amino acids including an extra copy of the reported immunodominant epitope DPYSPS. This study aimed to evaluate the IgE binding of the two Ara h 2 isoforms. Methods: Ten clones of Ara h 2 were sequenced to assess the relative frequency of the Ara h 2 isoforms and to identify whether there was further variation in the Ara h 2 sequence. IgE binding to Ara h 2.0101 and Ara h 2.0201 was measured for 70 peanut-allergic children using an IgE DELFIATM assay to quantitate specific IgE binding. A competition assay was used to measure whether Ara h 2.0201 contained IgE epitopes other than those found for Ara h 2.0101. Results: The original Ara h 2.0101 sequence was found for 6/10 clones and Ara h 2.0201 was found for 2/10 clones. Ara h 2.0201 had the expected insertion of 12 amino acids as well as substitutions at posit...
Clinical & Experimental Allergy, 2007
Background In peanut-allergic adults, IgE is mainly directed to Ara h1 and Ara h2. More recently, a role for Ara h6 has been suggested. In contrast to adults, IgE in children can fluctuate over time. Therefore, children may have a more dynamic reactivity to peanut. Objective To examine the IgE reactivity to major peanut allergens in peanut-allergic children at two subsequent time-points. Methods Twenty children (3-15 years old) with peanut allergy, confirmed by a double-blind placebo-controlled food challenge (DBPCFC), were included. Just before and 20 months after DBPCFC, IgE reactivity to purified Ara h1, Ara h2, Ara h3 and Ara h6 was studied by immunoblots and skin prick tests (SPTs). Results Before DBPCFC, all peanut-allergic children showed IgE reactivity to Ara h2; Ara h6 was recognized by 16 children, and Ara h1 and Ara h3 by 10 children. After 20 months, peanut-specific IgE levels (median 23 kU/L) and the individual recognition of major allergens were comparable with the levels and recognition before challenge (median 28.2 kU/L). SPT with Ara h2 and Ara h6 was positive in most children, whereas SPT with Ara h1 and Ara h3 was positive in approximately half of the children. Ara h6 induced the largest weals. None of the parameters were related to the severity of peanut allergy. Conclusion Ara h2 and Ara h6 are the most frequently recognized major peanut allergens in children. The individual reactivity to the major peanut allergens remained stable over time, despite DBPCFC.
Clinical & Experimental Allergy, 2005
Background IgE-binding peanut proteins smaller than 15 kDa were previously identified as potential allergens in the majority of our peanut allergic population. Objective To characterize the novel allergen in order to determine whether it was similar to one of the thus far identified recombinant peanut allergens (Ara h 1-7). Methods An IgE-binding protein of o15 kDa was purified and identified via N-terminal sequencing. Its IgE-binding properties were investigated using immunoblotting, basophil degranulation, and skin prick testing. Possible cross-reacting epitopes with other peanut allergens were studied using IgEimmunoblotting inhibition. Results The purified protein is a monomeric protein with a molecular weight of 14 981 Da as determined using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectroscopy. The amino acid sequence of the first 39 N-terminal residues is identical to that of Ara h 6, indicating that the allergen is Ara h 6. It is recognized by 20 out of 29 peanut-allergic patients on IgEimmunoblot, and its potent biological functionality is demonstrated by the degranulation of basophils, even at concentrations below 10 pg/mL, and by positive skin prick reactions. Ara h 6 has homology to Ara h 2, especially in the middle part and at the C-terminal part of the protein. Almost complete inhibition of IgE-Ara h 6 interaction with Ara h 2 demonstrates that at least part of the epitopes of Ara h 6 are cross-reactive with epitopes on Ara h 2. Conclusions Peanut-derived Ara h 6 is a biologically active allergen recognized by the majority of our peanut-allergic patient population and can be considered a clinically relevant peanut allergen.
IgE epitopes of Ara h 9, Jug r 3, and Pru p 3 in peanut-allergic individuals from Spain and the US
Frontiers in allergy, 2023
Non-specific lipid transfer proteins (LTPs) are well studied allergens that can lead to severe reactions, but often cause oral allergy syndrome in the Mediterranean area and other European countries. However, studies focused on LTP reactivity in allergic individuals from the United States are lacking because they are not considered major allergens. The goal of this study is to determine if differences in immunoglobulin (Ig) E binding patterns to the peanut allergen Ara h 9 and two homologous LTPs (walnut Jug r 3 and peach Pru p 3) between the US and Spain contribute to differences observed in allergic reactivity. Synthetic overlapping 15-amino acid-long peptides offset by five amino acids from Ara h 9, Jug r 3, and Pru p 3 were synthesized, and the intact proteins were attached to microarray slides. Sera from 55 peanut-allergic individuals from the US were tested for IgE binding to the linear peptides and IgE binding to intact proteins using immunofluorescence. For comparison, sera from 17 peanut-allergic individuals from Spain were also tested. Similar IgE binding profiles for Ara h 9, Jug r 3, and Pru p 3 were identified between the US and Spain, with slight differences. Certain regions of the proteins, specifically helices 1 and 2 and the C-terminal coil, were recognized by the majority of the sera more often than other regions of the proteins. While serum IgE from peanut-allergic individuals in the US binds to peptides of Ara h 9 and its homologs, only IgE from the Spanish subjects bound to the intact LTPs. This study identifies Ara h 9, Jug r 3, and Pru p 3 linear epitopes that were previously unidentified using sera from peanut-allergic individuals from the US and Spain. Certain regions of the LTPs are recognized more often in US subjects, indicating that they represent conserved and possible cross-reactive regions. The location of the epitopes in 3D structure models of the LTPs may predict the location of potential conformational epitopes bound by a majority of the Spanish patient sera. These findings are potentially important for development of peptide or protein-targeting diagnostic and therapeutic tools for food allergy.