Highly sensitive and wide-range quantification of bovine α_S1-casein in foods for predicting potential milk allergenicity using an IgE multi-epitope-specific antibody

Cow's milk allergy (CMA) is a common immunoglobulin E (IgE)-mediated food allergy, with α_S1-casein being the major allergen. Therefore, IgE epitopes could serve as reliable biomarkers for α_S1-casein detection and milk allergenicity prediction. Here, a conventional sandwich enzyme-linked immunosorbent assay (pAb-tα_S1-casein-based sELISA) and an Fe-N-C single-atom nanozymes (Fe-N-C SANs) probe-based sELISA were developed using a polyclonal antibody targeting nine IgE epitopes of α_S1-casein (pAb-tα_S1-casein) to detect α_S1-casein and predict potential milk allergenicity. The developed assays showed high sensitivity, specificity, accuracy, precision, recoveries, and ultra-wide linear detection ranges (spanning four to five orders of magnitude) for α_S1-casein detection. The capacity of these assays to predict milk allergenicity in real samples was confirmed using serum IgE from CMA patients. These results highlight the reliability of the developed methods for the highly sensitive quantification of α_S1-casein and the prediction of potential milk allergenicity in foods.