Novel Bead-Based Epitope Assay is a sensitive and reliable tool for profiling epitope-specific antibody repertoire in food allergy

• Published in: Scientific reports 2019-12, Vol.9 (1), p.18425-18425, Article 18425
• Main Authors: Suprun, Maria, Getts, Robert, Raghunathan, Rohit, Grishina, Galina, Witmer, Marc, Gimenez, Gustavo, Sampson, Hugh A., Suárez-Fariñas, Mayte
• Format: Article
• Language: English
• Subjects: 631/250/24/2510; 631/61/32; 82/1; 82/75; Adolescent; Adult; Allergens - immunology; Allergens - metabolism; Allergies; Animals; Arachis - immunology; Biological Variation, Individual; Child; Epitope Mapping - methods; Epitopes; Epitopes - immunology; Epitopes - metabolism; Female; Food; Food allergies; Food Hypersensitivity - diagnosis; Food Hypersensitivity - immunology; High-Throughput Screening Assays - methods; Humanities and Social Sciences; Humans; Immunoglobulin E; Immunoglobulin E - immunology; Immunoglobulin E - metabolism; Immunoglobulin G; Male; Milk - immunology; multidisciplinary; Peptide Library; Peptides; Phenotypes; Prognosis; Reproducibility of Results; Science; Science (multidisciplinary); Sensitivity and Specificity; Severity of Illness Index; Young Adult
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-019-54868-7

Identification of allergenic IgE epitopes is instrumental for the development of novel diagnostic and prognostic methods in food allergy. In this work, we present the quantification and validation of a Bead-Based Epitope Assay (BBEA) that through multiplexing of epitopes and multiple sample processing enables completion of large experiments in a short period of time, using minimal quantities of patients’ blood. Peptides that are uniquely coupled to beads are incubated with serum or plasma samples, and after a secondary fluorophore-labeled antibody is added, the level of fluorescence is quantified with a Luminex reader. The signal is then normalized and converted to epitope-specific antibody binding values. We show that the effect of technical artifacts, i.e. well position or reading order, is minimal; and batch effects - different individual microplate runs - can be easily estimated and eliminated from the data. Epitope-specific antibody binding quantified with BBEA is highly reliable, reproducible and has greater sensitivity of epitope detection compared to peptide microarrays. IgE directed at allergenic epitopes is a sensitive biomarker of food allergy and can be used to predict allergy severity and phenotypes; and quantification of the relationship between epitope-specific IgE and IgG4 can further improve our understanding of the immune mechanisms behind allergic sensitization.