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Background: The majority of children with cow's milk allergy (CMA) tolerate baked milk. However, reactivity to fermented milk products such as yogurt/cheese has not been previously evaluated. We sought to determine whether children with CMA could tolerate yogurt/cheese and whether a patient's IgE and IgG4‐binding pattern to milk protein epitopes could distinguish clinical reactivity. Methods: Four groups of reactivity were identified by Oral food challenge: baked milk reactive, fermented milk reactive, whole milk reactive, and outgrown. sIgE and sIgG4 binding to milk protein epitopes were assessed with a novel Luminex‐based peptide assay (LPA). Using machine learning techniques, a model was developed to predict different degrees of CMA. Results: The baked milk reactive patients demonstrated the highest degree of IgE epitope binding, which was followed sequentially by fermented milk reactive, whole milk reactive, and outgrown. Data were randomly divided into two groups with 75% of the data utilized for model development (n = 68) and 25% for testing (n = 21). All 68 children used for training were correctly classified with models using IgE and IgG4 epitopes. The average cross‐validation accuracy was much higher for models using IgE plus IgG4 epitopes by LPA (84.8%), twice the performance of the serum component proteins assayed by UniCAP (41.9%). The performance of the model on "unseen data" was tested using the 21 withheld patients, and the accuracy of IgE was 86% (AUC = 0.89) while of IgE+IgG4 model was 81% (AUC = 0.94). Conclusion: Using a novel high‐throughput LPA, we were able to distinguish the diversity of IgE/IgG4 binding to epitopes in the varying CMA phenotypes. LPA is a promising tool to predict correctly different degrees of CMA. The new food challenge sequence and Luminex‐based peptide assay to identify the clinical phenotypes of reactivity to baked, fermented, and whole cow's milk. [ABSTRACT FROM AUTHOR]

Background Oral immunotherapy ( OIT) with cow's milk ( CM) has been reported to induce a number of specific antibody responses, but these remain to be fully characterized. Our objective was to explore whether IgE and IgG4 epitope binding profiles could predict the risk of side effects during CM OIT. Methods The study population consisted of 32 children (6-17 yr of age) with CM allergy: 26 children who successfully completed OIT and six children who discontinued therapy due to adverse reactions. We investigated sera drawn before and after OIT. We analyzed specific IgE and IgG4 binding to CM protein-derived peptides with a microarray-based immunoassay. Antibody binding affinity was analyzed with a competition assay where CM proteins in solution competed with peptides printed on the microarray. Results IgE binding to CM peptides decreased and IgG4 binding increased following the OIT in children who attained desensitization. Compared with children who successfully completed OIT, those who discontinued OIT due to adverse reactions developed increased quantities and affinity of epitope-specific IgE antibodies and a broader diversity of IgE and IgG4 binding, but less overlap in IgE and IgG4 binding to CM peptides. Conclusions Detailed analysis of IgE and IgG4 binding to CM peptides may help in predicting whether CM OIT will be tolerated successfully. It may thus improve the safety of the therapy. [ABSTRACT FROM AUTHOR]

Background: Patients with peanut allergy have highly stable pathologic antibody repertoires to the immunodominant B-cell epitopes of the major peanut allergens Ara h 1 to 3. Objective: We used a peptide microarray technique to analyze the effect of treatment with peanut oral immunotherapy (OIT) on such repertoires. Methods: Measurements of total peanut-specific IgE (psIgE) and peanut-specific IgG4 (psIgG4) were made with CAP-FEIA. We analyzed sera from 22 patients with OIT and 6 control subjects and measured serum specific IgE and IgG4 binding to epitopes of Ara h 1 to 3 using a high-throughput peptide microarray technique. Antibody affinity was measured by using a competitive peptide microarray, as previously described. Results: At baseline, psIgE and psIgG4 diversity was similar between patients and control subjects, and there was broad variation in epitope recognition. After a median of 41 months of OIT, polyclonal psIgG4 levels increased from a median of 0.3 μg/mL (interquartile range [25% to 75%], 0.1-0.43 μg/mL) at baseline to 10.5 μg/mL (interquartile range [25% to 75%], 3.95-45.48 μg/mL; P < .0001) and included de novo specificities. psIgE levels were reduced from a median baseline of 85.45 kUA/L (23.05-101.0 kUA/L) to 7.75 kUA/L (2.58-30.55 kUA/L, P < .0001). Affinity was unaffected. Although the psIgE repertoire contracted in most OIT-treated patients, several subjects generated new IgE specificities, even as the total psIgE level decreased. Global epitope-specific shifts from IgE to IgG4 binding occurred, including at an informative epitope of Ara h 2. Conclusion: OIT differentially alters Ara h 1 to 3 binding patterns. These changes are variable between patients, are not observed in control subjects, and include a progressive polyclonal increase in IgG4 levels, with concurrent reduction in IgE amount and diversity. [Copyright &y& Elsevier]