Background: Certain immunotoxic peptides from gluten are resistant to gastrointestinal digestion and can interact with celiac-patient factors to trigger an immunologic response. A gluten-free diet (GFD) is the only effective treatment for celiac disease (CD), and its compliance should be monitored to avoid cumulative damage. However, practical methods to monitor diet compliance and to detect the origin of an outbreak of celiac clinical symptoms are not available.Objective: We assessed the capacity to determine the gluten ingestion and monitor GFD compliance in celiac patients by the detection of gluten and gliadin 33-mer equivalent peptidic epitopes (33EPs) in human feces.Design: Fecal samples were obtained from healthy subjects, celiac patients, and subjects with other intestinal pathologies with different diet conditions. Gluten and 33EPs were analyzed by using immunochromatography and competitive ELISA with a highly sensitive antigliadin 33-mer monoclonal antibody.Results: The resistance of a significant part of 33EPs to gastrointestinal digestion was shown in vitro and in vivo. We were able to detect gluten peptides in feces of healthy individuals after consumption of a normal gluten-containing diet, after consumption of a GFD combined with controlled ingestion of a fixed amount of gluten, and after ingestion of <100 mg gluten/d. These methods also allowed us to detect GFD infringement in CD patients.Conclusions: Gluten-derived peptides could be sensitively detected in human feces in positive correlation with the amount of gluten intake. These techniques may serve to show GFD compliance or infringement and be used in clinical research in strategies to eliminate gluten immunotoxic peptides during digestion. This trial was registered at clinicaltrials.gov as NCT01478867.

The GlutenTox® Pro Test is an immunochromatographic test for the detection of gluten in foods and on surfaces with varying compositions and levels of processing, from raw foods/ingredients to final product testing. The Method Developer evaluation for the validation of the GlutenTox Pro Test Kit (Biomedal Diagnostics, Sevilla, Spain) for the detection of gluten in foods and on surfaces was conducted at Biomedal, S. L., Camas, Sevilla, Spain. The GlutenTox Pro test method was evaluated by testing the following: cross-reactivity, interference, specificity and sensitivity, robustness, stability, lot-to-lot variation, food matrix, and environmental surface. To evaluate the performance of the GlutenToxPro test for the detection of gluten, 10 matrixes were selected: rice flour, bread/biscuit, rolled oat, pâté, and yogurt (and a second bread matrix for incurred sampled testing) for the food matrix study and food-grade painted wood, plastic, rubber, sealed ceramic, and stainless steel for the environmental surface matrix study. For the food matrix study, 30 replicates were evaluated at six spiked levels of gluten (0, 3, 8, 15, 25, and 45 ppm) against four detection thresholds (5, 10, 20, and 40 ppm) for each food matrix. Additionally, 10 replicates were evaluated at a concentration of 10,000 ppm using all four detection thresholds only for rice flour matrix. Three replicates of each concentration level of gluten were analyzed using paired samples by the AOAC OMA 2012.01 reference method for each food matrix. For the environmental surface study, 30 replicates were evaluated at a low spike level of gluten (16 ng/16 cm2), five replicates at a high spike level of gluten (400 ng/16 cm2), and five replicates at an unspiked control level (0 ng/16 cm2) for each surface matrix. Upon completion of testing, the probability of detection values and confidence intervals were calculated and plotted versus the concentration level as determined by the reference method when applicable. An independent laboratory evaluation of the GlutenTox Pro Test Kit with rice flour and stainless steel environmental surface was conducted at Q Laboratories, Inc. (Cincinnati, OH). The GlutenTox Pro Test Kit demonstrated reliability as an effective rapid method for the detection of gluten in food matrixes (LOD 5 ppm gluten; threshold limits 5, 10, 20, and 40 ppm gluten) and on environmental surfaces (amount of detection 16 ng/16 cm2).

Most gluten analysis methods have been developed to detect intact gluten, but they have shown limitations in certain foods and beverages in which gluten proteins are hydrolyzed. Methods based on G12/A1 moAbs detect the sequences of gluten immunogenic peptides (GIP), which are the main contributors to the immune response of celiac disease (CD). Immunogenic sequences with tandem epitopes for G12/A1 have been found in beers with <20 mg/kg gluten, which could be consumed by CD patients according to the Codex Alimentarius. Therefore, an accurate method for the estimation of the immunogenicity of a beer is to use two moAbs that can recognize celiac T cell epitopes comprising most of the immunogenic response. Here, a specific and sensitive method based on G12/A1 LFIA was developed to detect GIP in beers labeled gluten-free or with low gluten content, with an LOD of 0.5 mg/kg. A total of 107 beers were analyzed, of those 6.5% showed levels higher than 20 mg/kg gluten and 29% showed levels above the LOD. In addition, G12/A1 LFIA detected gluten in 15 more beer samples than competitive ELISA with another antibody. Despite their labeling, these beers contained GIP which may cause symptoms and/or intestinal damage in CD patients.

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