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Detection of wheat allergens using 2D Western blot and mass spectrometry.
J Pharm Biomed Anal 2019; 178:112907JP

Abstract

BACKGROUND

Wheat allergy is relatively common and the associated clinical manifestations depend on the involved molecular allergens as well as on the way of exposure. Different symptoms have been described: wheat-dependent exercise-induced anaphylaxis (WDEIA), atopic dermatitis (AD) and pollen rhinitis (PR). Traditional diagnostic methods do not allow accurate molecular identification of the allergens that are essential for risk assessment and for the choice of the most adapted treatment.

METHODS

Standardized total protein extracts obtained from wheat seeds were separated by 2D electrophoresis. Twenty-one sera with high wheat-specific immunoglobulin E (sIgE) levels were classified into three patients groups based on their clinical profile. These sera were tested by Western blot on 2D separated standardized wheat protein extract and their sIgE sensitization profiles were compared.

RESULTS

Specific sensitization profiles were identified for each phenotype group. For WDEIA, protein spots around 37 kDa (pH 6-9) and 37-50 kDa (pH 5-6) were identified. For AD, spots were observed around 50 kDa (pH 9), 10 kDa (pH 9) and 20 to 75 kDa (pH3). For PR, specific spots were located around 90 kDa (pH 9). The mass spectrometry (UHPLC-MS/MS) analysis of these identified spots pointed out several potential interesting allergens: Tri a 26, Tri a bA, Tri a 34, Tri a tritin.

CONCLUSIONS

The present study allowed the identification of different protein areas specific to these studied groups. The protein spots of interest were identified by UHPLC-MS/MS. It has been possible to establish a link between a specific symptomatology and the newly identified responsible allergens.

Authors+Show Affiliations

CRIG, Liège, Belgium; University of Liège, Liège, Belgium. Electronic address: j.courtois@crig.be.CHU of Liège, Liège, Belgium.CRIG, Liège, Belgium; HELMo, Liège, Belgium.Allergopôle, Clinique Saint-Luc, Bouge, Belgium.University of Liège, Liège, Belgium; CHU of Liège, Liège, Belgium.CRIG, Liège, Belgium; HELMo, Liège, Belgium.CER Group, Marloie, Belgium.CHU of Liège, Liège, Belgium.CRIG, Liège, Belgium; HELMo, Liège, Belgium.

Pub Type(s)

Journal Article
Review

Language

eng

PubMed ID

31634755

Citation

Courtois, J, et al. "Detection of Wheat Allergens Using 2D Western Blot and Mass Spectrometry." Journal of Pharmaceutical and Biomedical Analysis, vol. 178, 2019, p. 112907.
Courtois J, Bertholet C, Tollenaere S, et al. Detection of wheat allergens using 2D Western blot and mass spectrometry. J Pharm Biomed Anal. 2019;178:112907.
Courtois, J., Bertholet, C., Tollenaere, S., Van der Brempt, X., Cavalier, E., El Guendi, S., ... Quinting, B. (2019). Detection of wheat allergens using 2D Western blot and mass spectrometry. Journal of Pharmaceutical and Biomedical Analysis, 178, p. 112907. doi:10.1016/j.jpba.2019.112907.
Courtois J, et al. Detection of Wheat Allergens Using 2D Western Blot and Mass Spectrometry. J Pharm Biomed Anal. 2019 Oct 9;178:112907. PubMed PMID: 31634755.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Detection of wheat allergens using 2D Western blot and mass spectrometry. AU - Courtois,J, AU - Bertholet,C, AU - Tollenaere,S, AU - Van der Brempt,X, AU - Cavalier,E, AU - El Guendi,S, AU - Gillard,N, AU - Gadisseur,R, AU - Quinting,B, Y1 - 2019/10/09/ PY - 2019/04/23/received PY - 2019/09/20/revised PY - 2019/10/01/accepted PY - 2019/10/22/pubmed PY - 2019/10/22/medline PY - 2019/10/22/entrez KW - Mass spectrometry KW - Western blot KW - Wheat allergens KW - Wheat allergy SP - 112907 EP - 112907 JF - Journal of pharmaceutical and biomedical analysis JO - J Pharm Biomed Anal VL - 178 N2 - BACKGROUND: Wheat allergy is relatively common and the associated clinical manifestations depend on the involved molecular allergens as well as on the way of exposure. Different symptoms have been described: wheat-dependent exercise-induced anaphylaxis (WDEIA), atopic dermatitis (AD) and pollen rhinitis (PR). Traditional diagnostic methods do not allow accurate molecular identification of the allergens that are essential for risk assessment and for the choice of the most adapted treatment. METHODS: Standardized total protein extracts obtained from wheat seeds were separated by 2D electrophoresis. Twenty-one sera with high wheat-specific immunoglobulin E (sIgE) levels were classified into three patients groups based on their clinical profile. These sera were tested by Western blot on 2D separated standardized wheat protein extract and their sIgE sensitization profiles were compared. RESULTS: Specific sensitization profiles were identified for each phenotype group. For WDEIA, protein spots around 37 kDa (pH 6-9) and 37-50 kDa (pH 5-6) were identified. For AD, spots were observed around 50 kDa (pH 9), 10 kDa (pH 9) and 20 to 75 kDa (pH3). For PR, specific spots were located around 90 kDa (pH 9). The mass spectrometry (UHPLC-MS/MS) analysis of these identified spots pointed out several potential interesting allergens: Tri a 26, Tri a bA, Tri a 34, Tri a tritin. CONCLUSIONS: The present study allowed the identification of different protein areas specific to these studied groups. The protein spots of interest were identified by UHPLC-MS/MS. It has been possible to establish a link between a specific symptomatology and the newly identified responsible allergens. SN - 1873-264X UR - https://www.unboundmedicine.com/medline/citation/31634755/Detection_of_wheat_allergens_using_2D_Western_blot_and_mass_spectrometry L2 - https://linkinghub.elsevier.com/retrieve/pii/S0731-7085(19)30973-2 DB - PRIME DP - Unbound Medicine ER -