Ara h 2 and Ara 6 are the best predictors of severe peanut allergy: a double-blind placebo-controlled study.

BACKGROUND: Component-resolved diagnostics offers a modern tool in peanut allergy, but studies applying consistently double-blind placebo-controlled challenges are lacking. We aimed to optimize diagnostics for moderate-to-severe peanut allergy in a birch-endemic region and to create an oral-peanut challenge with its allergen activity characterized. METHODS: We performed double-blind placebo-controlled peanut challenges for a referred sample of 6- to 18-year-olds with peanut sensitization or a high suspicion of peanut allergy, including anaphylaxis. We measured specific IgE (sIgE) to Ara h 1, 2, 3, 6, 8, and 9. Testing of allergen activity of the challenge products was by IgE microarray inhibition. RESULTS: Of the 102 patients, 69 were challenge positive: 25 (36%) had severe, 36 (52%) moderate, and 8 (12%) mild symptoms; 38 (37%) received adrenalin. SIgE to Ara h 6 AUC 0.98 (95%CI, 0.96-1.00) was the best marker of moderate-to-severe allergy. When sIgE to Ara h 2 and Ara h 6 was measured together, all (100%) severe reactions at low doses were successfully diagnosable. SIgE to Ara h 8 had no diagnostic value, AUC 0.42 (95%CI, 0.30-0.52). Both nonroasted and roasted peanut inhibited 100% of IgE binding to Ara h 1, 2, 3, and 6. Nonroasted peanut inhibited 87% of IgE binding to Ara h 8, roasted inhibited 30%. The products lacked Ara h 9 activity. CONCLUSION: Co-sensitization to Ara h 2 and Ara h 6 was associated with severe reactions distinguishing severe allergy from mild symptoms. SIgE to Ara h 8 added no diagnostic value. Component-resolved diagnostics reduce the need for oral challenges in peanut allergy.

Invariant NKT cells are more abundant in peanut-allergic adults and a subset of CD8+ iNKT cells are depleted after peanut oil exposure.

Introduction: Peanut allergy is one of the most prevalent food allergies globally. Currently, most research into the mechanisms involved in protein allergy focuses on the protein allergens under investigation, and information on the function of accompanying compounds, such as lipids, is scarce. Thus, this research investigates the role of peanut-associated lipids and invariant natural killer T (iNKT) cells in peanut allergy using a novel, human, in vitro assay. Methods: PBMCs from non-allergic and peanut-allergic subjects were stimulated with the glycolipid, α-Galactosylceramide (α-GalCer), over 14 days for iNKT cell expansion. Autologous dendritic cells (DCs) were stimulated with either peanut oil, the lipid-binding peanut allergen, Ara h 8, or both peanut oil and Ara h 8. The expanded iNKT cells were then immunomagnetically isolated and co-cultured for 5 h with autologous DCs, and cytokine expression was measured by flow cytometry. Results: A 5-fold higher iNKT cell population was observed in peanut-allergic subject peripheral blood compared to non-allergic controls. In all subjects, conventional flow analysis highlighted iNKTs co-cultured with autologous α-GalCer-pulsed DCs displayed increased IL-4 and IFN-y secretion within 5 hours of co-culture. A 10-parameter unsupervised clustering analysis of iNKT phenotype found significantly more CD3+CD8+CD25+IL-4+IL-5+IL-10+IFNγ+ cells in non-allergic adults following culture with peanut oil. Conclusion: For the first time, we show iNKT cells are more abundant in peanut-allergic adults compared to non-allergic adults, and peanut lipid-exposed iNKT cells resulted in the identification of a subset of CD8+ iNKT cells which was significantly lower in peanut-allergic adults. Thus, this study proposes a role for iNKT cells and peanut allergen-associated lipids in peanut allergy.

An Improved Enzyme-Linked Immunosorbent Assay (ELISA) Based Protocol Using Seeds for Detection of Five Major Peanut Allergens Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8.

Peanut allergy is an important health concern among many individuals. As there is no effective treatment to peanut allergy, continuous monitoring of peanut-based products, and their sources is essential. Precise detection of peanut allergens is key for identification and development of improved peanut varieties with minimum or no allergens in addition to estimating the levels in peanut-based products available in food chain. The antibody based ELISA protocol along with sample preparation was standardized for Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8 to estimate their quantities in peanut seeds. Three different dilutions were optimized to precisely quantify target allergen proteins in peanut seeds such as Ara h 1 (1/1,000, 1/2,000, and 1/4,000), Ara h 2 and Ara h 3 (1/5,000, 1/10,000, and 1/20,000), Ara h 6 (1/40,000, 1/80,000, and 1/1,60,000), and Ara h 8 (1/10, 1/20, and 1/40). These dilutions were finalized for each allergen based on the accuracy of detection by achieving <20% coefficient of variation in three technical replicates. This protocol captured wide variation of allergen proteins in selected peanut genotypes for Ara h 1 (77-46,106 µg/g), Ara h 2 (265-5,426 µg/g), Ara h 3 (382-12,676 µg/g), Ara h 6 (949-43,375 µg/g), and Ara h 8 (0.385-6 µg/g). The assay is sensitive and reliable in precise detection of five major peanut allergens in seeds. Deployment of such protocol allows screening of large scale germplasm and breeding lines while developing peanut varieties with minimum allergenicity to ensure food safety.

Hypoallergen Peanut Lines Identified Through Large-Scale Phenotyping of Global Diversity Panel: Providing Hope Toward Addressing One of the Major Global Food Safety Concerns.

Peanut allergy is one of the serious health concern and affects more than 1% of the world's population mainly in Americas, Australia, and Europe. Peanut allergy is sometimes life-threatening and adversely affect the life quality of allergic individuals and their families. Consumption of hypoallergen peanuts is the best solution, however, not much effort has been made in this direction for identifying or developing hypoallergen peanut varieties. A highly diverse peanut germplasm panel was phenotyped using a recently developed monoclonal antibody-based ELISA protocol to quantify five major allergens. Results revealed a wide phenotypic variation for all the five allergens studied i.e., Ara h 1 (4-36,833 µg/g), Ara h 2 (41-77,041 µg/g), Ara h 3 (22-106,765 µg/g), Ara h 6 (829-103,892 µg/g), and Ara h 8 (0.01-70.12 µg/g). The hypoallergen peanut genotypes with low levels of allergen proteins for Ara h 1 (4 µg/g), Ara h 2 (41 µg/g), Ara h 3 (22 µg/g), Ara h 6 (829 µg/g), and Ara h 8 (0.01 µg/g) have paved the way for their use in breeding and genomics studies. In addition, these hypoallergen peanut genotypes are available for use in cultivation and industry, thus opened up new vistas for fighting against peanut allergy problem across the world.

Peanut allergens: new consolidated findings on structure, characteristics, and allergome.

Immunoglobulin E-mediated food allergy is the result of a complex pathomechanism. Factors contributing to the dysfunction of the immune system are the allergenic sources and the variable matrix effects arising from the processes involved in interaction with the gastrointestinal tract, the allergens themselves through their structural features, and the specific behavior of the individual immune system. The starting point for elucidating the pathomechanism of food allergy is the identification of allergens and the description of their structure. They are the basis for in vitro diagnostics as well as the development of immunotherapeutic drugs. With regard to Class I food allergy, peanut allergy affects by far the largest group of patients. 11 allergens have been identified in peanuts. Ara h 1, Ara h 3, and Ara h 4 belong to the cupin superfamily, Ara h 2, Ara h 6, and Ara h 7 to the prolamin superfamily; Ara h 5 (profilins) and Ara h 8 (superfamily of Bet v 1-homologous proteins) are associated with aeroallergens. Peanut lipid transfer proteins (LTP) and two peanut oleosins are listed as Ara h 9, Ara h 10, and Ara h 11 by the IUIS Allergen Nomenclature Subcommittee. Peanut agglutinin (PNA) and a third oleosin have been shown to possess allergenic properties. The effect of the above specified allergens has to be considered in the context of their matrix, which is influenced by processing factors.

Peanut sensitization pattern in Norwegian children and adults with specific IgE to peanut show age related differences.

BACKGROUND: Peanuts contain potent food allergens and the prevalence of allergy is reported to increase, especially in children. Since peanut sensitization may differ between different geographical regions, we wanted to investigate the sensitization pattern to the individual peanut allergens in a Norwegian population. METHODS: Cases reported to the Norwegian Food Allergy Register with sera positive to peanut extract were analyzed for specific IgE (sIgE) to the recombinant peanut allergens Ara h 1, Ara h 2, Ara h 3, Ara h 8 and Ara h 9 and to birch pollen extract. Serum samples negative to the above allergens were analyzed for sIgE to Ara h 6, and sIgE to Pru p 3 in peach were analyzed in sera positive to the cross-reactive allergen Ara h 9. RESULTS: Highest frequency of sIgE to Ara h 2, often co-sensitized to Ara h 1 and 3, were found in the small children up to 6 years of age. From the age of 6 years, sensitization to Ara h 8 was predominant. The sIgE levels to the storage proteins Ara h 1, 2 and 3 were strongly correlated, as was the sIgE levels to Ara h 8 and birch pollen extract. A low sensitization rate of sIgE to Ara h 9 in young adults was observed, which sIgE levels were very strongly correlated to Pru p 3. CONCLUSION: The sensitization to peanut allergens in a Norwegian population shows a clear age dependent pattern. The results add to the previously published research on the sensitization patterns of peanut sensitized patients in different geographical areas.

The value of specific IgE to peanut and its component Ara h 2 in the diagnosis of peanut allergy.

BACKGROUND: To avoid unnecessary oral food challenges, which are time consuming, stressful, and risky, improved in vitro diagnostic methods for food allergy such as component resolved diagnostics are still under investigation. OBJECTIVE: To investigate the role of whole peanut- and peanut-component (Ara h 1, Ara h 2, Ara h 3, Ara h 6 and Ara h 8)-specific IgE levels in the diagnostic procedure of peanut allergy as well as the diagnostic properties of peanut-specific IgG and IgG4. METHODS: Sixty-one children underwent oral peanut challenge tests for diagnostic purposes irrespective of their peanut-specific IgE levels. Peanut-specific serum IgE, IgG, and IgG4 levels were determined by ImmunoCAP FEIA and specific IgE against individual peanut proteins by Immuno Solid-phase Allergen Chip. RESULTS: Thirty-four of 61 patients (56%) had a peanut allergy. No significant difference was observed for peanut-specific IgG or peanut-specific IgG4 levels between patients who were allergic and tolerant patients, whereas peanut-specific IgE was significant higher in patients who were allergic than in tolerant patients (P < .005). Twenty-five of 61 children had peanut-specific IgE above a previously proposed cutoff level of 15 kUA/L; however, 7 of these 25 children (28%) were clinically tolerant. Ara h 2-specific IgE was significantly lower in tolerant than in patients with allergies (P < .0001). Interestingly, 94% of the patients with peanut allergies showed IgE-binding to Ara h 2. Unfortunately, 26% of the sensitized but tolerant patients have shown IgE binding to Ara h 2 too. CONCLUSIONS: Neither the level of specific IgE to peanut nor to Ara h 2 was able to clearly distinguish patients with clinical relevant peanut allergy from those who were clinical tolerant in our population. As expected, peanut-specific IgG and IgG4 did not improve the diagnostic procedure.

The utility of peanut components in the diagnosis of IgE-mediated peanut allergy among distinct populations.

BACKGROUND: Increasing data suggest that analysis of IgE to peanut components can be clinically helpful and possibly more accurate than IgE to whole peanut. Not all studies examining this topic, however, have used prospective samples, multiple components, and peanut challenges. OBJECTIVE: We sought to determine the utility of peanut component testing, using a standardized, commercially available test done before oral peanut challenge in various populations of patients with suspected peanut allergy from 2 different countries. METHODS: IgE to whole peanut and the recombinant allergen components Ara h 1, 2, 3, and 8 were analyzed from serum samples drawn before double-blind peanut challenge from 4 distinct cohorts of patients with suspected peanut allergy from 2 nations (United States and Sweden). RESULTS: Patients (n = 167; median age, 11.7 years; interquartile range, 7.0-15.0 years) had serum analyzed for peanut components and completed an oral food challenge to peanut. Although IgE to peanut was the most sensitive test (0.93), Ara h 2 was the most specific (0.92) and provided the best positive predictive value (0.94) of all the tests. Ara h 2 was also the best overall diagnostic test by receiver operating characteristic analysis (area under the curve, 0.84; P < .05). CONCLUSIONS: In patients with suspected peanut allergy, IgE to peanut is a sensitive test but is not specific. IgE to Ara h 2 is a more specific and more accurate diagnostic test in this sampling of patients with suspected peanut allergy. Given each tests attributes, a stepwise approach to testing may provide clinicians with a way to minimize the need for peanut challenges.