The Validation of the Wheat Gluten ELISA Kit

Abstract

Background: It is important to analyze the presence of wheat/gluten in food to avoid wheat allergy or celiac disease. Objective: The Wheat/Gluten ELISA kit was developed to measure total wheat protein or gluten content in wheat, barley, and rye cereals as raw materials, and processed foods. Validation as to whether this kit is suitable for quantifying total wheat protein/gluten was carried out. Methods: The Wheat/Gluten ELISA kit was designed as a sandwich ELISA based on antigliadin polyclonal antibody. Selectivity, interference study, matrix study including incurred food, robustness, stability, and lot-to-lot consistency studies were conducted for the Wheat/Gluten ELISA kit. Incurred matrix studies were also conducted in an independent laboratory. Results: The analysis of 38 different substances revealed no cross-reactivity above the LOQ except for oats. Recoveries of the spiked samples were mostly in the range of 75–140%, including an independent laboratory result. The LOD of the ELISA was found to be 0.02–0.16 mg/kg. Robustness testing proved that extraction time and incubation time of first reaction and enzyme reaction had no significant influence on quantified value. The stability at 2–8°C was found to exceed 12 months. Good lot-to-lot consistency was observed. Conclusions: The Wheat/Gluten ELISA kit showed good analytical performance in the quantitative analysis of total wheat protein/gluten in the identified food products using the AOAC Performance Tested Method(s)SM program. Highlights: The Wheat/Gluten ELISA kit was validated and showed good analytical performance in the quantitative analysis of total wheat protein/gluten in food.

A sensitive sandwich enzyme‐linked immunosorbent assay (sELISA) targeted multiple wheat protein fractions for the detection of several cereal grains in processed foods

“…Wheat, barley, and rye are taxonomically classified into the Gramineae family by phylogenetic relationships; thus, the close taxonomy and evolutionary relationship may have a greater chance of causing cross‐reactivity due to the similarity in protein and protein structure (Jones et al., 1995). Similar cross‐reactivity to barley and rye ingredient was also observed in other fabricated wheat or gluten ELISA methods (Koerner et al., 2013; Saito et al., 2009). A competitive ELISA for the quantification of gluten using a monoclonal antibody specific for the DQ2.5‐glia‐α3 T cell epitope from α‐gliadins (Sajic et al., 2017).…”

“…Wheat, barley, and rye are taxonomically classified into the Gramineae family by phylogenetic relationships; thus, the close taxonomy and evolutionary relationship may have a greater chance of causing cross-reactivity due to the similarity in protein and protein structure (Jones et al, 1995). Similar crossreactivity to barley and rye ingredient was also observed in other fabricated wheat or gluten ELISA methods (Koerner et al, 2013;Saito et al, 2009). A competitive ELISA for the F I G U R E 2 Extraction efficiency and developed wheat sELISA detectability analysis of the wheat protein extracts using standard sample preparation procedure from the wheat flour that has been moist heated and dry heated at different temperatures for 10 min.…”

Rapid, Effective, and Versatile Extraction of Gluten in Food with Application on Different Immunological Methods

“…It is necessary to optimize the extraction systems to produce the complete release and recovery of both prolamins and glutelins [46,47]. The time and temperature of incubation during the extraction process are two critical factors to consider [48].…”

Challenges in Gluten Analysis: A Comparison of Four Commercial Sandwich ELISA Kits

“…The success of gluten-free labeling relies on the accuracy and limitations of the analytical methods used to detect gluten residues. Multiple, quantitative commercial methods, primarily enzyme-linked immunosorbent assays (ELISAs), are available for the detection of gluten residues in foods [ 18 , 19 , 20 ]. ELISA-based methods are relatively simple, fast, and have become the most widely used gluten detection methods.…”