Comparison of R5 and G12 Antibody-Based ELISA Used for the Determination of the Gluten Content in Official Food Samples

Abstract

Celiac Disease (CD) is one of the most common food intolerances. It comes along with serious damage of the mucosa in the small intestine and is caused by the storage proteins—termed “gluten”—of wheat, rye, barley and possibly oats. Sensitive individuals need to stick to a strict gluten-free diet. The gluten level in food products labeled as “gluten-free”, must not exceed 20 mg/kg. It is obvious that effective test methods are needed to accurately determine the gluten concentration in foods. The determination of the presence of gluten in foodstuffs is mainly done by means of an immunochemical method called ELISA (enzyme-linked immunosorbent assay). To check the suitability of a G12 antibody-based gluten detection kit for its use in official control systems a number of routine samples were tested in parallel with two different test kits, as would be done in a routine lab. The determination of the gluten content was performed on samples entering the official laboratory including samples from official control plans, commercially available and private samples to request gluten-free labels. The results obtained with the G12 antibody ELISA assay were comparable to the official R5 method. A validation of the two different methods was not part of this study.

Gluten Analysis of Wheat Starches with Seven Commercial ELISA Test Kits—Up to Six Different Values

“…At the moment, the R5 Mendez method is recommended as type 1 method for the determination of gluten (Codex Standard 234-1999, but due to recent progress in method development and validation studies (Don et al 2014), efforts are underway to review this recommendation and add the G12 mAb-based method or others (Codex Committee of Methods of Analysis and Sampling 2015). A recent comparison of results obtained with the G12 compared to the R5 ELISA concluded that both assays gave comparable gluten contents in a variety of routine food samples (Hochegger et al 2015). The aforementioned study did not include any WSt samples, but the data presented here indicated that the R5 and G12 assays yielded significantly different gluten contents in 23 out of a total of 31 WSt, including the control sample (W9 + 20).…”

Immunological Methods in Gluten Risk Analysis: A Snapshot

“…Other research groups have recently compared the performance of the different antibodies [56][57][58][59] on several food matrices, from grains of different species (e.g., wheat, rye and barley) to fermented-hydrolyzed foods.…”

“…It is a shared position within the scientific community that the identification of the most suitable sample preparation and extraction procedure is one more critical point in gluten determination [32,33,59]. It is known that the main requirement for immunological assays is, in fact, that they should be able to measure the toxic proteins, regardless of the type of food (raw material or heat-treated and enzymatically-treated product).…”

“…It is known that the main requirement for immunological assays is, in fact, that they should be able to measure the toxic proteins, regardless of the type of food (raw material or heat-treated and enzymatically-treated product). The solvent most commonly used to extract the prolamins fraction from raw materials, such as flours, is aqueous alcohol, namely, 60% ethanol or 50% propanol [32,33,59]. Partially hydrolised gluten is soluble in aqueous alcohol solutions and also in water or salt solutions.…”

“…In heat-processed foods like bread, extruded products like pasta, or products containing hydrolysed gluten, proteins are degraded and hydrolyzed, or protein aggregates may form. This means that the aqueous alcohol solution is inadequate and the food toxicity can be underestimated [32,33,59]. To counteract these critical points, when gluten detection is made on thermally processed samples, additional agents (disaggregating and reducing) are used to break the interchain S-S-bonds of proteins and solubilise gluten.…”