Abstract
Background and AimsCeliac disease is a permanent intolerance to gluten prolamins from wheat, barley, rye and, in some patients, oats. Partially digested gluten peptides produced in the digestive tract cause inflammation of the small intestine. High throughput, immune-based assays using monoclonal antibodies specific for these immunotoxic peptides would facilitate their detection in food and enable monitoring of their enzymatic detoxification. Two monoclonal antibodies, G12 and A1, were developed against a highly immunotoxic 33-mer peptide. The potential of each antibody for quantifying food toxicity for celiac patients was studied.MethodsEpitope preferences of G12 and A1 antibodies were determined by ELISA with gluten-derived peptide variants of recombinant, synthetic or enzymatic origin.ResultsThe recognition sequences of G12 and A1 antibodies were hexameric and heptameric epitopes, respectively. Although G12 affinity for the 33-mer was superior to A1, the sensitivity for gluten detection was higher for A1. This observation correlated to the higher number of A1 epitopes found in prolamins than G12 epitopes. Activation of T cell from gluten digested by glutenases decreased equivalently to the detection of intact peptides by A1 antibody. Peptide recognition of A1 included gliadin peptides involved in the both the adaptive and innate immunological response in celiac disease.ConclusionsThe sensitivity and epitope preferences of the A1 antibody resulted to be useful to detect gluten relevant peptides to infer the potential toxicity of food for celiac patients as well as to monitor peptide modifications by transglutaminase 2 or glutenases.
Highlights
Celiac disease (CD) is a common autoimmune disorder that has genetic, environmental, and immunological components
To test for monoclonal antibodies (moAb) specificity, we studied the cross-reactivity values (CR) of these moAb against commercial gliadin, by indirect ELISA
The G12 moAb presented an IC50 of almost double that obtained with the A1 moAb, suggesting that A1 had broader reactivity with gliadin epitopes than G12, which is more specific for the 33-mer (Figure 1B)
Summary
Celiac disease (CD) is a common autoimmune disorder that has genetic, environmental, and immunological components. The ingestion of gluten proteins contained in wheat, barley, and rye, and, in some cases, oats [6,7], leads to characteristic inflammation, villous atrophy, and crypt hyperplasia in the CD patient upper small intestine [2]. The principal toxic components belong to a family of closely related proline and glutamine rich proteins called gliadins [8]. Celiac disease is a permanent intolerance to gluten prolamins from wheat, barley, rye and, in some patients, oats. Immune-based assays using monoclonal antibodies specific for these immunotoxic peptides would facilitate their detection in food and enable monitoring of their enzymatic detoxification. The potential of each antibody for quantifying food toxicity for celiac patients was studied
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