Introduction: Molecular studies of hen’s egg allergens help define allergic phenotypes, with IgE to sequential (linear) epitopes on the ovomucoid (OVM) protein associated with a persistent disease. Epitope profiles of other egg allergens are largely unknown. The objective of this study was to construct an epitope library spanning across 7 allergens and further evaluate sequential epitope-specific (ses-)IgE and ses- IgG4 among baked-egg reactive or tolerant children. Methods: A Bead-Based Epitope Assay was used to identify informative IgE epitopes from 15-mer overlapping peptides covering the entire OVM and ovalbumin (OVA) proteins in 38 egg allergic children. An amalgamation of 12 B-cell epitope prediction tools was developed using experimentally identified epitopes. This ensemble was used to predict epitopes from ovotransferrin, lysozyme, serum albumin, vitellogenin-II fragment, and vitellogenin-1 precursor. Ses- IgE and ses- IgG4 repertoires of 135 egg allergic children (82 reactive to baked-egg, the remaining 52 tolerant), 46 atopic controls, and 11 healthy subjects were compared. Results: 183 peptides from OVM and OVA were screened and used to create an aggregate algorithm, improving predictions of 12 individual tools. A final library of 65 sequential epitopes from 7 proteins was constructed. Egg allergic children had higher ses- IgE and lower ses- IgG4 to predominantly OVM epitopes than both atopic and healthy controls. Baked-egg reactive children had similar ses- IgG4 but greater ses- IgE than tolerant group. A combination of OVA-sIgE with ses- IgEs to OVM-023 and OVA-028 was the best predictor of reactive phenotype. Conclusion: We have created a comprehensive epitope library and showed that ses- IgE is a potential biomarker of baked-egg reactivity.

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