Allergen Assay Research Articles

Multiplex Specific IgE Profiling in Neonatal Stool of Preterms Predicts IgE-Mediated Disease

Background: The natural history of immunoglobulin (Ig) E-mediated diseases in preterm infants is still elusive. We aimed at developing a non-invasive tool for detecting specific IgE (sIgE) and eosinophil-derived neurotoxin (EDN) in neonatal fecal samples and evaluating its predictive value for the development of IgE-mediated diseases during the first year of life. Methods: We developed a stool extraction protocol, followed by freeze-drying and solubilization. The sIgEs were investigated in neonatal fecal samples from 21 preterm infants with a 300-allergen multiplex and confirmed by a capillary Western blot with a nano-immunoassay. EDN concentration was used to investigate the local eosinophilic component. Results: The multiplexed allergen assay detected sIgE in all of the samples. A Western blot was used to confirm the results. The frequency and levels of sIgE in the neonatal fecal samples differed between the infants who developed IgE-mediated diseases and the controls. Allergen specificity was associated with the development of cow’s milk allergy (CMA) and asthma. The development of CMA was predicted by the sIgE response to milk proteins (sensitivity was 88%; specificity was 78%). The EDN levels predicted the development of IgE-mediated diseases (sensitivity was 100%; specificity was 75%). Conclusion: The non-invasive investigation of neonatal fecal sIgE is a promising tool for predicting the subsequent development of IgE-mediated diseases. Clinical Implications: The non-invasive sIgE and EDN profiling of neonatal fecal samples from preterm infants can predict the development of IgE-mediated diseases.

High-stable perovskite nanocrystal fluorescent probe-based aptasensor for ultrasensitive detection of peanut allergen Ara h1

Food allergy is a serious immune allergy reaction, that has attracted public attention all over the world due to the increasing incidence. Ara h1, the most abundant allergenic protein in peanuts, has been identified as a biomarker of peanut allergen. Therefore, a sensitive, efficient method for Ara h1 detection is urgently needed to protect allergy sufferers. In this study, a simple and efficient fluorescent aptasensor was proposed for ultrasensitive and rapid detection of peanut allergen Ara h1. The aptamer against Ara h1 was conjugated onto the surface of high-stable perovskite nanocrystal (PNCs) to serve as the fluorescence-labeled recognition probe as well as the signal probe, while the magnetic nanoparticles Fe3O4 (MNPs) were functionalized with the aptamer complementary DNA (cDNA) as the separation probe. The high-stable PNCs-labeled aptasensor Apt-PNCs@cDNA-MNPs gives the combined merits of the extraordinary optical properties of PNCs, the efficient magnetic separation capability of MNPs as well as the high specificity of aptamers. The developed high-stable PNCs-labeled aptasensor exhibited excellent linearity (0.1–100 ng/mL), and low limit of detection (0.04 ng/mL). The aptasensor was successfully applied for the determination of Ara h1 in food samples, implying great potential in the field of allergen assay.

Optimised multiplex droplet digital PCR is more precise, but not more sensitive, than real-time PCR for the detection of allergenic peanut

The United States requires labelling of food products containing major allergens, such as peanut, through the Food Allergen Labeling and Consumer Protection Act. Accurate labelling requires sensitive, specific and robust detection methods, and PCR-based techniques have proven highly effective. This article describes the transition of a previously developed multiplex real-time PCR assay for allergenic peanut to a droplet digital PCR format. The triplex droplet digital PCR assay was developed in a probe mixing format and directly compared to the established real-time PCR assay. Data are provided for thorough optimisation in the digital format, including the effects of primer and probe concentration, cycle number and annealing/extension time. Optimisation parameters influenced relative location and separation of droplet clusters but not final copy number. The droplet digital PCR assay was linear over five orders of magnitude; its lower limit of detection was 0.05 pg DNA per reaction, more sensitive than published digital PCR allergen assays. It was more precise, but not more sensitive, than the previously established real-time PCR assay.

Evaluation of sensitization to Der p 1 and Der p 2 in a pediatric population of the North of Portugal

In Portugal, data on the role of Der p 1 and Der p 2 in patients with house dust mite (HDM) allergy are scarce. Allergen-specific immunotherapy (sIT) is the only treatment that improves symptoms, reduces the need for pharmacological therapy and modifies the natural history of the disease. With this study, the authors aim to understand the local epidemiology and to clarify if the molecular assay of major allergens is advantageous in deciding and/or modifying the decision to initiate sIT in children with clinical indication which are sensitized to Dermatophagoides pteronyssinus. Retrospective study with analysis of patients with asthma and/or rhinitis. January/2013-December/2016. 1) positive prick-test to Dermatophagoides pteronyssinus; and 2) clinically relevant disease under treatment. Assay Der p 1 and Der p 2 values ≥0.35 kUA/L were considered positive. Statistical significance was set at P<.05. The clinical files of 279 patients. Mean ages 9.55 years (min.4-max.17). Asthma was present in 199 children (71.3%) and rhinitis in 245 (87.8%). Der p 1 and Der p 2 was <0.35kUA/L in 29 (10,4%) patients. The value of Der p 1/Der p 2 correlated with the size of the prick-test papule, the value of the eosinophils and the total IgE. Der p 1 and Der p 2 are dominant allergens in our population and there may be benefits in determining these molecular allergen levels in patients with a positive prick-test and a clinical indication for sIT prior to a decision of initiating sIT or not.

Component-resolved diagnostics in vernal conjunctivitis.

Conventional diagnostic tests in allergy are insufficient to clarify the cause of vernal conjunctivitis. Component-resolved diagnostic (CRD) by microarray allergen assay may be useful in detecting allergens that might be involved in the inflammatory process. In a recent trial in patients suffered from eosinophilic esophagitis, after 2 years of the CRD-guided exclusion diet and specific immunotherapy, significant clinical improvement was observed, and 68% of patients were discharged (cure based on negative biopsy, no symptoms, and no medication intake). Our new objective was to evaluate IgE-mediated hypersensitivity by CRD in tears and serum from patients with vernal conjunctivitis and treat patients with identified triggering allergens by specific immunotherapy. Twenty-five patients with vernal conjunctivitis were evaluated. The identified triggering allergens were n Lol p 1 (11 cases), n Cyn d 1 (eight cases), group 4 and 6 grasses (six cases) and group 5 of grasses (five cases). Prick test and pollen IgE were positive in one case. Clinical improvement was observed in 13/25 vernal conjunctivitis patients after 1-year specific immunotherapy. CRD seems to be a more sensitive diagnostic tool compared with prick test and IgE detection. Specific CRD-led immunotherapy may achieve clinical improvements in vernal conjunctivitis patients.

Improved recombinant Api m 1- and Ves v 5-based IgE testing to dissect bee and yellow jacketallergy and their correlation with the severity of the sting reaction.

No study has assessed the diagnostic sensitivity of rApi m 1 and rVes v 5 on Immulite testing system. To compare the diagnostic sensitivity of commercially available venom recombinant allergens between the currently available immunoassays [ImmunoCAP (CAP) and Immulite (LITE)] and establish their correlation with the severity of the sting reaction. This study evaluated 95 bee venom and 110 yellow jacket venom-allergic subjects. We measured the levels of sIgE to rApi m 1, rVes v 5 (LITE and CAP), rApi m 2 (LITE), rVes v 1 (CAP) and total IgE (CAP). Forty-nine healthy subjects served as controls. The diagnostic sensitivity of rApi m 1 and rVes v 5 was significantly higher with the LITE than with the CAP system (71% vs. 88% and 82% vs. 93%). The specificity of both assays for both allergens was between 94% and 98%. Twenty-nine patients that tested negative for rApi m 1 or rVes v 5 with CAP were positive with LITE, but none of the patients that tested negative with LITE were positive with CAP. The positive values of rApi m 1 and rVes v 5 were on average 2.7 and 2.3 times higher, with the LITE than with the CAP system. The combination of rApi m 1 and rApi m 2 (LITE) and the combination of rVes v 5 (LITE) and rVes v 1 (CAP) almost matched the sensitivity of native venoms (95% and 97%, respectively), whereas the diagnostic sensitivity of the combination of rVes v 5 and rVes v 1 (CAP) did not reach the sensitivity of rVes v 5 (LITE) alone (90% vs. 93%). IgE levels to venom recombinants and total IgE did not correlate with the severity of sting reaction. The use of rApi m 1 and rVes v 5 with the LITE system significantly enhanced diagnostic utility of venom recombinants and should improve the dissection of bee and yellow jacket venom allergy.

Food Allergies: The Basics

IgE-associated food allergy affects approximately 3% of the population and has severe effects on the daily life of patients—manifestations occur not only in the gastrointestinal tract but also affect other organ systems. Birth cohort studies have shown that allergic sensitization to food allergens develops early in childhood. Mechanisms of pathogenesis include cross-linking of mast cell– and basophil-bound IgE and immediate release of inflammatory mediators, as well as late-phase and chronic allergic inflammation, resulting from T-cell, basophil, and eosinophil activation. Researchers have begun to characterize the molecular features of food allergens and have developed chip-based assays for multiple allergens. These have provided information about cross-reactivity among different sources of food allergens, identified disease-causing food allergens, and helped us to estimate the severity and types of allergic reactions in patients. Importantly, learning about the structure of disease-causing food allergens has allowed researchers to engineer synthetic and recombinant vaccines.

Quantitative Multiplex Real-Time PCR Assay for Shrimp Allergen: Comparison of Commercial Master Mixes and PCR Platforms in Rapid Cycling

Real-time PCR has been used widely in numerous fields. In food safety, it has been applied to detection of microbes and other contaminants, including food allergens. Interest in rapid (fast) cycling real-time PCR has grown because it yields results in less time than does conventional cycling. However, fast cycling can adversely affect assay performance. Here we report on tests of commercial master mixes specifically designed for fast real-time PCR using a shrimp allergen assay we previously developed and validated. The objective of this work was to determine whether specialized commercial master mixes lead to improved assay performance in rapid cycling. Real-time PCR assays were carried out using four different master mixes and two different rapid cycling protocols. Results indicated that specialized master mixes did yield quality results. In many cases, linear ranges spanned up to 7 orders of magnitude, R2 values were at least 0.95, and reaction efficiencies were within or near the optimal range of 90 to 110%. In the faster of the two rapid cycling protocols tested, assay performance and PCR amplification were markedly better for the shorter PCR product. In conclusion, specialized commercial master mixes were effective as part of rapid cycling protocols, but conventional cycling as used in our previous work is more reliable for the shrimp assay tested.

Value of microarray allergen assay in the management of eosinophilic oesophagitis

BackgroundEosinophilic oesophagitis (EoE) is a disorder characterised by oesophageal dysfunction and, histologically, by eosinophilic inflammation. Although treatment, which includes dilatations, oral corticosteroids and restrictive diets, is often effective, choosing the foods to be eliminated from the diet is difficult. ObjectiveComponent resolved diagnostic by microarray allergen assay may be useful in detecting allergens that might be involved in the inflammatory process. MethodsWe studied 67 patients with EoE, diagnosed clinically and histologically by endoscopic biopsy. CRD analysis with microarray technology was carried out in the 67 EoE patients, 50 patients with pollen allergy without digestive symptoms, and 50 healthy controls. ResultsAllergies were not detected by microarray in only seven of the 67 patients with EoE. Controls with pollen allergy showed sensitisation to different groups of pollen proteins without significant differences. In EoE patients with response to some allergens, the predominant allergens were grasses group 1 and, in particular, nCyn d 1 (Cynodon dactylon) or Bermuda grass pollen in 59.5%, followed by lipid transfer proteins (LTP) of peach (19.40%), hazelnut (17.91%) and Artemisia (19.40%). ConclusionsIn patients with EoE, sensitisation to plant foods and pollen is important. The proteins most frequently involved are nCyn d 1 and lipid transfer proteins, hazelnuts and walnuts. After one year of an array-guided exclusion diet and pollen-specific immunotherapy in the case of high levels of response, patients with EoE showed preliminary significant improvements.

The additional values of microarray allergen assay in the management of polysensitized patients with respiratory allergy

The IgE response is directed against specific components from an allergenic source. The traditional diagnostic methods use whole extracts, containing allergenic, nonallergenic and cross-reactive molecules. This may pose diagnostic challenges in polysensitized patients. Microarray techniques detect specific IgE against multiple molecules, but their value in term of additional information and economic saving has not been yet defined. We assessed the additional diagnostic information provided by an allergen microarray in a large population of polysensitized subjects. In this multicentre study, allergists were required to carefully record diagnosis and treatment of consecutive patients referred for asthma/rhinitis, using the standard methodology (history, skin prick test, IgE assay). Then, a microarray allergen assay was carried out. Clinicians were required to review their diagnosis/treatment according to microarray results. 318 allergic patients (30% reporting also nonrespiratory symptoms) and 91 controls were enrolled. The clinicians reported at least one additional information from the microarray in about 60% of patients, this resulting in therapeutic adjustments. In 66% of patients IgE to pan-allergens were detectable, being this clinically relevant in 38% of patients with polysensitization to pollens. Microarray IgE assay represents an advancement in allergy diagnosis, as a third-level approach in polysensitized subjects, when the traditional diagnosis may be problematic.

A personalized food allergen testing platform on a cellphone.

We demonstrate a personalized food allergen testing platform, termed iTube, running on a cellphone that images and automatically analyses colorimetric assays performed in test tubes toward sensitive and specific detection of allergens in food samples. This cost-effective and compact iTube attachment, weighing approximately 40 grams, is mechanically installed on the existing camera unit of a cellphone, where the test and control tubes are inserted from the side and are vertically illuminated by two separate light-emitting-diodes. The illumination light is absorbed by the allergen assay, which is activated within the tubes, causing an intensity change in the acquired images by the cellphone camera. These transmission images of the sample and control tubes are digitally processed within 1 s using a smart application running on the same cellphone for detection and quantification of allergen contamination in food products. We evaluated the performance of this cellphone-based iTube platform using different types of commercially available cookies, where the existence of peanuts was accurately quantified after a sample preparation and incubation time of ~20 min per test. This automated and cost-effective personalized food allergen testing tool running on cellphones can also permit uploading of test results to secure servers to create personal and/or public spatio-temporal allergen maps, which can be useful for public health in various settings.

A multi-center ring trial of allergen analysis using fluorescent multiplex array technology

BackgroundConsistent performance of allergen assays is essential to ensure reproducibility of exposure assessments for investigations of asthma and occupational allergic disease. This study evaluated intra- and inter-laboratory reproducibility of a fluorescent multiplex array, which simultaneously measures eight indoor allergens in a single reaction well. MethodsA multi-center study was performed in nine laboratories in the US and Europe to determine the inter-laboratory variability of an 8-plex array for dust mite, cat, dog, rat, mouse and cockroach allergens. Aliquots of 151 dust extract samples were sent to participating centers and analyzed by each laboratory on three separate occasions. Agreement within and between laboratories was calculated by the concordance correlation coefficient (CCC). ResultsResults were obtained for over 32,000 individual allergen measurements. Levels covered a wide range for all allergens from below the lower limit of detection (LLOD=0.1–9.8ng/ml) to higher than 6800ng/ml for all allergens except Mus m 1, which was up to 1700ng/ml. Results were reproducible within as well as between laboratories. Within laboratories, 94% of CCC were ≥0.90, and 80% of intra-laboratory results fell within a 10% coefficient of variance (CV%). Results between laboratories also showed highly significant positive correlations for all allergens (~0.95, p<0.001). Overall means of results were comparable, and inter-laboratory CV% for all allergens except Rat n 1 ranged between 17.6% and 26.6%. ConclusionThe data indicate that performance criteria for fluorescent multiplex array technology are reproducible within and between laboratories. Multiplex technology provides standardized and consistent allergen measurements that will streamline environmental exposure assessments in allergic disease.

Proteomic allergen–peptide/protein interaction assay for the identification of human skin sensitizers

Modification of proteins by skin sensitizers is a pivotal step in T cell mediated allergic contact dermatitis (ACD). In this process small reactive chemicals interact covalently or non-covalently with cellular or extracellular skin self-proteins or self-peptides to become recognized by the human immune system. Aiming to develop a novel non-animal in vitro test system for predicting sensitization potential of small reactive chemicals in human skin the allergen–peptide/protein interaction assay (APIA) has been developed. By applying modern proteomic technologies together with a target peptide containing all amino acids, the assay permits the profiling of all amino acid specific allergen–peptide interactions. Moreover, potentially crucial allergen-specific Cys-modifications are qualitatively monitored by mass spectrometry and confirmed by a dual peptide approach. Assay conditions chosen mimic the distinct human epidermal reactivity compartments of the skin surface (pH 5.5), stratum basale (pH 6.8), and typical physiological conditions (pH 7.4). An extreme as well as a moderate human contact sensitizer produced Cys-specific mass shifts, whereas a skin irritant did not. Our data indicate that MALDI–MS based and skin-related in vitro technology platforms – like the APIA – are promising tools in developing alternative non-animal allergen assays. This will assist in chemical classification and next generation risk assessment strategies, including REACH and experimental immunotoxicology.

Analysis of allergic sieving detection results and clinical significance in children with allergic disease

To explore the clinical use of sieving detection among the childhood with allergic disease. The sieving detection about allergen inhalant allergens, Fx5 in the CAP anaphylactogen detection system, and serum specific IgE were detected in three hundred and thirty-one cases of children (aged from 1 year to 14 years old) with allergic disease. Patients were divided into group 1, group 2 and group 3 according to the age from 0 to 3, 3 to 6, and 6 to 14 years old. All datas were statistical analysed among different age groups. Among the 331 patients, the positive rate of allergic sieving detection was 67.98%, the elevation rate of IgE was 53.78%. Inhalant allergen positive rate was 60.42%, while the food allergen positive rate was 28.10%. Inhalant allergen positive rate of the group 3 (aged from 6 to 14 years old) was significant higher than the other two age groups (68.45%). And the food allergen positive rate of the age group 1 (aged from 0 to 3 years old) was significant higher than the other two age groups (62.50%). Positive rate for simply inhalant allergen was 39.88%, while positive rate for simply food allergen was 7.55% and mixed allergen was 20.54%. Inhalant allergen was the main allergen of the children with allergic disease aged over 3 years old, while food allergen was the main allergen of the children with allergic disease aged below 3 years old. It was safe, sensible and effective to use Uni CAP anaphylactogen detection system for rapid assay of specific allergens.

50 The Allergic March Resolved at Allergen Component Level

BackgroundThe allergic march is well known at the level of pattern of sensitisation, but there is no information of its evolution in term of sensitzation to single allergenic molecules. We investigated the evolution of the IgE repertoire by means of a microarray allergen assay.MethodsSerum samples from allergic patients of a wide age range were analyzed by a micorarray chip, which allow to identify in a single assay the presence of specific IgE towards 103 allergenic molecules. Total IgE were also evaluated as an internal control. Patients were stratified in 6 groups according to their age (0–2; 3–5; 6–9; 10–13; 14–17 and >17 years).ResultsSamples from 609 patients were analysed. The behaviour of total IgE according to age strictly paralleled that of the sum of specific IgE. Food-related components were the more frequently recognized in the first ages, whereas specific IgE to plant allergens appeared later. Nonetheless, mite-specific IgE were the most represented in all age classes. Specific IgE against cross-reacting allergens were virtually absent in the first years and tended to appear after the age of 6.ConclusionsThe molecular pattern of allergen recognition according to age well reflects the clinical characteristics of the allergic march.

Adverse Food Reactions—An Emerging Issue for Adults

Adverse reactions to foods are classified according to the presence or absence of involvement of the immune system, which may or may not include the production of immunoglobulin E (IgE) antibodies. This review focuses on the epidemiology, diagnosis, and management of adverse food reactions, primarily in adults, and excluding celiac disease and lactose intolerance. Reported reactions to foods are often believed to be manifestations of a food allergy; however, IgE-mediated food allergy only affects 1% to 4% of adults, with seafood, tree nuts, peanuts, fruits, and vegetables being the most common triggers. Diagnosis is challenging and most commonly achieved through careful evaluation of clinical history followed by elimination and reintroduction or challenge with the suspected offending food. With acute-onset allergic reactions, estimation of food-specific IgE antibodies is frequently used to confirm or refute the diagnosis. Recent developments, such as single allergen assays, enhance the diagnosis of IgE-mediated food allergy, but the gold standard remains oral food challenge. Despite recent advances in the management of food allergy, including the promotion of oral tolerance, the mainstay of management is still the avoidance of food triggers. Dietary management can be compromised by nutritional inadequacy, accidental exposure, food labeling, and quality of life or adherence issues. It is essential that adults with confirmed food allergy receive optimal nutrition and dietetic support to enable them to manage their condition.

Production et standardisation des extraits allergéniques en 2011

The aim of the standardization of allergen products for diagnosis and immunotherapy of allergy is to deliver batches, qualitatively and quantitatively reproducible. As allergens are extracted from natural sources (pollen, mites, molds, danders, venoms, food), the objective of the production is to reflect the complexity of the allergens present in the source materials. The biological standardization is based on comparison of each production batch with a qualified In-House Reference (IHR) by IgE-binding assays. The IHR is formerly calibrated by in vivo by skin prick-test on sensitized patients. Major allergen assays are also developed to measure relevant molecular allergens in the different batches. French and European regulation issues have been recently updated, according to the recent evolution of the scientific knowledge about allergens. Homologous groups have been defined, clarifying the set of pharmaceutical data needed for the representative allergen of a group, and therefore for non-grouped allergens, used for diagnosis and immunotherapy of allergy.

The History of IgE: From Discovery to 2010

The discovery of IgE and the role of IgE-mediated inflammation gave clinical allergy a scientific backbone, and as a result, the reputation of the specialty allergy has increased considerably over the years. Allergy diagnosis was improved by assays for in vitro determination of the presence and concentration of IgE antibodies, and clinical knowledge also broadened, allowing better service for the increasing number of allergic individuals. Access to immune assays for allergens finally allowed characterization and standardization of allergen preparations used for diagnosis and allergen-specific immunotherapy. Improved basic molecular technologies have further increased our knowledge about the complex component composition of an allergen extract, introduced IgE-specific immunotherapy, and allowed the allergist to better handle even severe allergic reactions such as anaphylaxis.

Stability and mixing compatibility of dog epithelia and dog dander allergens

Little information or data are available concerning the stability and compatibility of dog epithelia and dog dander allergens. To determine the immunochemical reactivities of commercial, nonstandardized dog epithelia and dog dander extracts after exposures to various temperatures or after mixing with high-protease fungal and cockroach extracts at concentrations recommended for maintenance immunotherapy (IT) injections. Quantitative enzyme-linked immunosorbent assay and qualitative (immunoblot) analyses were performed to compare specific compositional changes with total or individual allergen activities. Assays for dog allergens Can f 1 and Can f 3 (albumin) used specific mouse or rabbit antibodies. Multiallergen enzyme-linked immunosorbent assay inhibition and immunoblot methods were conducted using a human serum pool with high levels of IgE to dog allergens. Dog allergen recoveries ranged from 22% to 134% after short exposures to moderate or extreme temperatures and from 28% to 118% after mixing with fungal or insect extracts and storage for up to 15 months at 2 degrees C to 8 degrees C. Recoveries in dog dander extracts varied up to 2.5-fold with different test methods. Immunoblots revealed partial degradation of dog albumin molecules to discrete fragments that retained antibody-binding activities. In most cases, recoveries improved at elevated glycerin concentrations. Dog allergens in epithelia and dander extracts exhibited favorable temperature stabilities. Compatibilities with fungal or insect extracts may be compromised or at risk in some combinations. These data support current IT practice parameter recommendations of separating high-protease extracts from other products if possible; they also demonstrate that dog extracts possess allergen stabilities suitable for many IT formulations.

Hypoallergenic Characteristics of Wheat Flour Produced by Stepwise Polishing

Hypoallergenic wheat flour was produced by polished-graded method using a rice polishing machine. Eight fractions (C1–C8) of polished-graded wheat flours were obtained step wise from the outer layer of whole wheat grains by 10% of the total weight, and the distribution of allergenic protein in each fraction was determined. The salt-soluble (albumin/globulin), salt-insoluble (glutenin) and alcohol-soluble (gliadin) proteins obtained from the polished-graded wheat flours were tested for the allergen assay with immunodetection using the sera of wheat allergenic patients. Immunoblotting results confirmed that the innermost fraction (C8) contained a smaller amount of allergenic proteins. Albumin/globulin groups in all fractions (C1–C8) showed different IgE-reactivity patterns, the 60–75 kDa proteins appeared in all of fraction flours. It was higher in C3–C5 fractions. Fractions C3 and C4 contained higher amount of specific wheat allergenic protein including 60–75, 35, 22, and < 20 kDa. IgE-antibody also bound to glutenin (25–37 kDa), especially in C3 and C4, but the binding proteins showed quite faint bands. Gliadin was found in all fractions of wheat flour. We propose that polishing is an appropriate method to obtain hypoallergenic wheat flours, and the fraction C8 may be possible to be consumed by people suffering from wheat allergy.