A sensitive sandwich enzyme-linked immunosorbent assay (sELISA) for the detection of walnut residues in processed food

Walnut can trigger IgE-mediated food allergy and seriously endangers life quality and health of walnut allergy patients. The objective of the study was to develop a specific and sensitive sandwich enzyme-linked immunosorbent assay (sELISA) to detect multiple walnut protein fractions and explore the effects of processing and food matrix on total protein dissolution and ELISA detection in order to assure accurate labeling and consumer protection. The detection limit (LOD) was 7.38 ng/mL and the recovery in food matrices ranged in between 52.92% and 111.97%, whereas the intra- and inter-assay coefficients of variation were less than 10% and 15%, respectively. The developed sELISA was highly specific with cross-reactivity of less than 1% with coix seed, maize, hazelnut, pistachio, cashew and tomato proteins among almost 29 foods and food ingredients tested. This method was preliminarily applied for the detection of commercial food, and the detection results were basically consistent with the label indications. Furthermore, baking above 180 ℃ resulted in a significant reduction in protein extraction and ELISA recovery. Thermal processing causing a greater reduction of recovery in the assay than non-thermal processing, with dry thermal processing having a greater impact than wet thermal processing. In model foods, the effects of processing and food matrices resulted in lower level of proteins extraction with corresponding reduced detectability. These findings indicated that the sELISA developed in this research possess a great significance on the rapid and accurate detection of walnut protein.