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Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System

The detection of gluten in foodstuffs has become a growing concern in food allergen management as a result of the high ratio of population sensitive to the main gluten-containing cereals. In this study, a promising single-domain antibody previously isolated by phage display (dAb8E) was produced in P...

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Autores principales: García-García, Aina, Madrid, Raquel, Garcia-Calvo, Eduardo, Mendoza-Chamizo, Belén, García, Teresa, Martin, Rosario
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7764234/
https://www.ncbi.nlm.nih.gov/pubmed/33321826
http://dx.doi.org/10.3390/foods9121838
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author García-García, Aina
Madrid, Raquel
Garcia-Calvo, Eduardo
Mendoza-Chamizo, Belén
García, Teresa
Martin, Rosario
author_facet García-García, Aina
Madrid, Raquel
Garcia-Calvo, Eduardo
Mendoza-Chamizo, Belén
García, Teresa
Martin, Rosario
author_sort García-García, Aina
collection PubMed
description The detection of gluten in foodstuffs has become a growing concern in food allergen management as a result of the high ratio of population sensitive to the main gluten-containing cereals. In this study, a promising single-domain antibody previously isolated by phage display (dAb8E) was produced in Pichia pastoris resulting in high levels of the antibody fragment expression (330 mg/L). The purified dAb8E was proved to specifically bind to gluten proteins from wheat, barley and rye, exhibiting no cross reaction to other heterologous species. The dynamic range of the sandwich enzyme-linked immunosorbent assay (ELISA) covered 0.1 to 10 µg/mL of gliadin, reaching a limit of detection of 0.12 µg/mL. When experimental binary mixtures of the target cereals were analyzed, the limit of detection was 0.13 mg/g, which would theoretically correspond to gluten concentrations of approximately 13 mg/kg. Finally, thirty commercially available food products were analyzed by means of the developed assay to further confirm the applicability of the dAb8E for gluten determination. The proposed methodology enabled the generation of a new gluten-specific nanobody which could be used to guarantee the appropriate labelling of gluten-free foods.
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spelling pubmed-77642342020-12-27 Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System García-García, Aina Madrid, Raquel Garcia-Calvo, Eduardo Mendoza-Chamizo, Belén García, Teresa Martin, Rosario Foods Article The detection of gluten in foodstuffs has become a growing concern in food allergen management as a result of the high ratio of population sensitive to the main gluten-containing cereals. In this study, a promising single-domain antibody previously isolated by phage display (dAb8E) was produced in Pichia pastoris resulting in high levels of the antibody fragment expression (330 mg/L). The purified dAb8E was proved to specifically bind to gluten proteins from wheat, barley and rye, exhibiting no cross reaction to other heterologous species. The dynamic range of the sandwich enzyme-linked immunosorbent assay (ELISA) covered 0.1 to 10 µg/mL of gliadin, reaching a limit of detection of 0.12 µg/mL. When experimental binary mixtures of the target cereals were analyzed, the limit of detection was 0.13 mg/g, which would theoretically correspond to gluten concentrations of approximately 13 mg/kg. Finally, thirty commercially available food products were analyzed by means of the developed assay to further confirm the applicability of the dAb8E for gluten determination. The proposed methodology enabled the generation of a new gluten-specific nanobody which could be used to guarantee the appropriate labelling of gluten-free foods. MDPI 2020-12-10 /pmc/articles/PMC7764234/ /pubmed/33321826 http://dx.doi.org/10.3390/foods9121838 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
García-García, Aina
Madrid, Raquel
Garcia-Calvo, Eduardo
Mendoza-Chamizo, Belén
García, Teresa
Martin, Rosario
Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title_full Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title_fullStr Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title_full_unstemmed Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title_short Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System
title_sort production of a recombinant single-domain antibody for gluten detection in foods using the pichia pastoris expression system
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7764234/
https://www.ncbi.nlm.nih.gov/pubmed/33321826
http://dx.doi.org/10.3390/foods9121838
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