Sensitive detection of cereal fractions that are toxic to celiac disease patients by using monoclonal antibodies to a main immunogenic wheat peptide.
BACKGROUND
Celiac disease is an immune-mediated enteropathy caused by the ingestion of gluten, a protein fraction found in certain cereals. Immunotoxic gluten peptides that are recalcitrant to degradation of digestive enzymes appear to trigger celiac syndromes. A 33-mer peptide from alpha-2 gliadin has been identified as a principal contributor to gluten immunotoxicity. A gluten-free diet is the usual first therapy for celiac disease patients; therefore, the characterization and quantification of the toxic portion of the gluten in foodstuffs is crucial to avoid celiac damage.


OBJECTIVE
We aimed to develop immunologic assays as a novel food analysis tool for measuring cereal fractions that are immunotoxic to celiac disease patients.


METHODS
The design focused on the production of monoclonal antibodies against the gliadin 33-mer peptide and the development of enzyme-linked immunosorbent assays (ELISAs) and Western blot analysis with the use of novel antibodies.


RESULTS
A sandwich ELISA method showed a detection limit for wheat, barley, and rye of <1 ppm prolamine. However, the method required a sample that was > or =1 order of magnitude greater for the detection of low-toxic oats, and there was no signal with the safe cereals maize and rice. A competitive ELISA method was also developed for detection of the toxic peptide in hydrolyzed food, which had a detection limit of <0.5 ppm gliadin.


CONCLUSIONS
Both ELISAs designed for use with the toxic gliadin 33-mer peptide suggested a high correlation between the presence of the peptide and the amount of cereal that was toxic to celiac disease patients. The sensitivity was significantly higher than that of equivalent methods recognizing other gluten epitopes.
Morón
Belén
B
Departamento de Microbiología y Parasitología, Facultad de Farmacia, Universidad de Sevilla, Seville, Spain.
Cebolla
Angel
A
Manyani
Hamid
H
Alvarez-Maqueda
Moisés
M
Megías
Manuel
M
Thomas
María Del Carmen
Mdel C
López
Manuel Carlos
MC
Sousa
Carolina
C
D000911:Antibodies, Monoclonal; D018554:Avena / Q000276:immunology; D002446:Celiac Disease / Q000139:chemically induced* / Q000276:immunology; D002523:Edible Grain / Q000737:chemistry* / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D000939:Epitopes; D005504:Food Analysis / Q000379:methods*; D005903:Gliadin / Q000032:analysis* / Q000633:toxicity*; D001467:Hordeum / Q000276:immunology; D006801:Humans; D012275:Oryza / Q000276:immunology; D010446:Peptide Fragments / Q000032:analysis / Q000633:toxicity; D011994:Recombinant Proteins; D012434:Secale / Q000276:immunology; D012680:Sensitivity and Specificity; D014908:Triticum / Q000276:immunology; D003313:Zea mays / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010446:Peptide Fragments; D011994:Recombinant Proteins; D005903:Gliadin