Haoran Gao1, Yining Jin1, Dan Ioan Jian1, Eric Olson2, Perry K W Ng3, Venu Gangur4. 1. Food Allergy & Immunology Laboratory, Michigan State University, East Lansing, MI 48824, United States. 2. Wheat Breeding and Genetics Laboratory, Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI 48824, United States. 3. Cereal Science Laboratory, Department of Food Science & Human Nutrition, Michigan State University, East Lansing, MI 48824, United States. 4. Food Allergy & Immunology Laboratory, Michigan State University, East Lansing, MI 48824, United States. Electronic address: gangur@msu.edu.
Abstract
BACKGROUND: Wheat allergy is a major food allergy that has reached significant levels of global public health concern. Potential variation in allergenicity among different wheat genotypes is not well studied at present largely due to the unavailability of validated methods. Here, we developed and validated a novel mouse-based primary screening method for this purpose. METHODS: Groups of Balb/c mice weaned on-to a plant protein-free diet were sensitized with salt-soluble protein (SSP) extracted from AABB genotype of wheat (durum, Carpio variety). After confirming clinical sensitization for anaphylaxis, mice were boosted 7 times over a 6-month period. Using a pooled-plasma mini bank, a wheat-specific IgE-inhibition (II)-ELISA was optimized. Then the relative allergenicity of SSPs from tetraploid (AABB), hexaploid (AABBDD) and diploid (DD) wheat genotypes were determined. The IC50/IC75 values were estimated using IgE inhibition curves. RESULTS: The optimized II-ELISA with an inhibition time of 2.5 h had a co-efficient of variation of <2%. Primary screening for relative allergenicity demonstrated that IgE binding to AABB-SSP was significantly abolished by the other two wheat genotypes. Compared to AABB, the relative allergenicity of SSPs of AABBDD and DD were significantly lower (p < .01). Furthermore, IgE inhibition curves showed significant differences in IC50 and IC75 values among the three wheat genotypes. CONCLUSION: We report a novel mouse-based primary screening method of testing relative allergenicity of wheat proteins from three different wheat genotypes for the first time. This method is expected to have broad applications in wheat allergy research.
BACKGROUND:Wheatallergy is a major food allergy that has reached significant levels of global public health concern. Potential variation in allergenicity among different wheat genotypes is not well studied at present largely due to the unavailability of validated methods. Here, we developed and validated a novel mouse-based primary screening method for this purpose. METHODS: Groups of Balb/c mice weaned on-to a plant protein-free diet were sensitized with salt-soluble protein (SSP) extracted from AABB genotype of wheat (durum, Carpio variety). After confirming clinical sensitization for anaphylaxis, mice were boosted 7 times over a 6-month period. Using a pooled-plasma mini bank, a wheat-specific IgE-inhibition (II)-ELISA was optimized. Then the relative allergenicity of SSPs from tetraploid (AABB), hexaploid (AABBDD) and diploid (DD) wheat genotypes were determined. The IC50/IC75 values were estimated using IgE inhibition curves. RESULTS: The optimized II-ELISA with an inhibition time of 2.5 h had a co-efficient of variation of <2%. Primary screening for relative allergenicity demonstrated that IgE binding to AABB-SSP was significantly abolished by the other two wheat genotypes. Compared to AABB, the relative allergenicity of SSPs of AABBDD and DD were significantly lower (p < .01). Furthermore, IgE inhibition curves showed significant differences in IC50 and IC75 values among the three wheat genotypes. CONCLUSION: We report a novel mouse-based primary screening method of testing relative allergenicity of wheat proteins from three different wheat genotypes for the first time. This method is expected to have broad applications in wheatallergy research.
Authors: Yining Jin; Haoran Gao; Rick Jorgensen; Jillian Salloum; Dan Ioan Jian; Perry K W Ng; Venugopal Gangur Journal: Int J Mol Sci Date: 2020-05-01 Impact factor: 5.923
Authors: Rick Jorgensen; Rajsri Raghunath; Haoran Gao; Eric Olson; Perry K W Ng; Venu Gangur Journal: Int J Mol Sci Date: 2022-06-10 Impact factor: 6.208