Tags

Type your tag names separated by a space and hit enter

Innovative approach to low-level gluten determination in foods using a novel sandwich enzyme-linked immunosorbent assay protocol.
Eur J Gastroenterol Hepatol. 2003 May; 15(5):465-74.EJ

Abstract

OBJECTIVES

There is currently much call for a reliable enzyme-linked immunosorbent assay (ELISA) protocol for determining gluten in foods to serve as a basis for further Codex Alimentarius regulations. Given its ability to recognize the potential coeliac-toxic epitope QQPFP, which occurs repeatedly in alpha-, gamma- and omega-gliadins, hordeins and secalins, the monoclonal antibody R5 raised against a secalin extract may prove to be an essential tool for gluten analysis. This study was designed to develop a highly sensitive and specific sandwich ELISA to quantify low levels of wheat, barley and rye prolamins in foods for coeliacs.

METHODS

Simple sandwich ELISA based on the use of a single monoclonal antibody (R5) as both the coating and detection was developed. A quantitative cocktail gluten-extraction procedure for heat-processed foods was also tested.

RESULTS

R5-ELISA was able to identify gliadins, hordeins and secalins with assay sensitivities of 0.78, 0.39 and 0.39 ng/ml, respectively. The assay's detection limit was 1.5 ng gliadins/ml (1.56 ppm gliadins, 3.2 ppm gluten). The system proved insensitive to the non-coeliac-toxic cereals maize, rice and oats, and was non-cultivar-dependent. It was also able to detect gliadins and hordeins in unprocessed and heat-processed wheat- and barley-based products, and to estimate the gluten content of hydrolysed foods.

CONCLUSION

We present a new generation of a robust sandwich R5-ELISA with good reproducibility (8.7%) and repeatability (7.7%). Its gluten-detection limit of 3.2 ppm is lower than the existing threshold of 20-200 ppm. The ELISA, which is equally sensitive to barley, wheat and rye prolamins, is compatible with the quantitative cocktail extraction procedure for heat-processed foods. Along with the cocktail procedure, the Working Group on Prolamin Analysis and Toxicity is currently evaluating an R5-ELISA system as proposed by the Codex Alimentarius Commission.

Authors+Show Affiliations

Unidad de Gluten, Centro Nacional de Biotecnología, Madrid, Spain.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

12702901

Citation

Valdés, Israel, et al. "Innovative Approach to Low-level Gluten Determination in Foods Using a Novel Sandwich Enzyme-linked Immunosorbent Assay Protocol." European Journal of Gastroenterology & Hepatology, vol. 15, no. 5, 2003, pp. 465-74.
Valdés I, García E, Llorente M, et al. Innovative approach to low-level gluten determination in foods using a novel sandwich enzyme-linked immunosorbent assay protocol. Eur J Gastroenterol Hepatol. 2003;15(5):465-74.
Valdés, I., García, E., Llorente, M., & Méndez, E. (2003). Innovative approach to low-level gluten determination in foods using a novel sandwich enzyme-linked immunosorbent assay protocol. European Journal of Gastroenterology & Hepatology, 15(5), 465-74.
Valdés I, et al. Innovative Approach to Low-level Gluten Determination in Foods Using a Novel Sandwich Enzyme-linked Immunosorbent Assay Protocol. Eur J Gastroenterol Hepatol. 2003;15(5):465-74. PubMed PMID: 12702901.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Innovative approach to low-level gluten determination in foods using a novel sandwich enzyme-linked immunosorbent assay protocol. AU - Valdés,Israel, AU - García,Enrique, AU - Llorente,Mercedes, AU - Méndez,Enrique, PY - 2003/4/19/pubmed PY - 2003/7/15/medline PY - 2003/4/19/entrez SP - 465 EP - 74 JF - European journal of gastroenterology & hepatology JO - Eur J Gastroenterol Hepatol VL - 15 IS - 5 N2 - OBJECTIVES: There is currently much call for a reliable enzyme-linked immunosorbent assay (ELISA) protocol for determining gluten in foods to serve as a basis for further Codex Alimentarius regulations. Given its ability to recognize the potential coeliac-toxic epitope QQPFP, which occurs repeatedly in alpha-, gamma- and omega-gliadins, hordeins and secalins, the monoclonal antibody R5 raised against a secalin extract may prove to be an essential tool for gluten analysis. This study was designed to develop a highly sensitive and specific sandwich ELISA to quantify low levels of wheat, barley and rye prolamins in foods for coeliacs. METHODS: Simple sandwich ELISA based on the use of a single monoclonal antibody (R5) as both the coating and detection was developed. A quantitative cocktail gluten-extraction procedure for heat-processed foods was also tested. RESULTS: R5-ELISA was able to identify gliadins, hordeins and secalins with assay sensitivities of 0.78, 0.39 and 0.39 ng/ml, respectively. The assay's detection limit was 1.5 ng gliadins/ml (1.56 ppm gliadins, 3.2 ppm gluten). The system proved insensitive to the non-coeliac-toxic cereals maize, rice and oats, and was non-cultivar-dependent. It was also able to detect gliadins and hordeins in unprocessed and heat-processed wheat- and barley-based products, and to estimate the gluten content of hydrolysed foods. CONCLUSION: We present a new generation of a robust sandwich R5-ELISA with good reproducibility (8.7%) and repeatability (7.7%). Its gluten-detection limit of 3.2 ppm is lower than the existing threshold of 20-200 ppm. The ELISA, which is equally sensitive to barley, wheat and rye prolamins, is compatible with the quantitative cocktail extraction procedure for heat-processed foods. Along with the cocktail procedure, the Working Group on Prolamin Analysis and Toxicity is currently evaluating an R5-ELISA system as proposed by the Codex Alimentarius Commission. SN - 0954-691X UR - https://www.unboundmedicine.com/medline/citation/12702901/Innovative_approach_to_low_level_gluten_determination_in_foods_using_a_novel_sandwich_enzyme_linked_immunosorbent_assay_protocol_ L2 - http://dx.doi.org/10.1097/01.meg.0000059119.41030.df DB - PRIME DP - Unbound Medicine ER -