Pediatrics Resident Lucile Packard Children's Hospital Stanford Mountain View, California, United States
Background: Peanut allergy affects more than 1 in every 50 children. The best predictor of clinical reactivity is an elevated serum IgE titer against peanut proteins Ara h2 or 6. Yet, among patients with high peanut-specific serum IgE, only a subset will have clinically significant reactions to oral challenge. Previous research has characterized peanut-specific IgE in allergic subjects, but how these antibodies differ in sensitized but tolerant subjects and how these differences contribute to allergic reactions remain incompletely understood. Objective: I will compare B cell populations and receptor sequences in 20 subjects with peanut allergy, 20 subjects with peanut sensitization but tolerance, and 5 nonallergic, nonsensitized controls. I hypothesize that allergic subjects, as compared to sensitized, tolerant subjects, will have significantly higher frequencies of circulating peanut-specific B cells and plasmablasts and that the peanut allergen epitopes targeted by B cells will differ between these cohorts. This research will provide insight into the contribution of peanut-specific B cells to clinical reactions and may provide direction for designing therapeutic antibodies and immunotherapies. Design/Methods: Biobanked samples were collected under institutional IRB approval. To identify peanut-specific B cell populations, I will immunophenotype and isolate antigen-specific B cells by single-cell flow sorting using allergen tetramer constructs labeled with fluorophores and DNA barcodes. Then, I will amplify and sequence single-cell transcriptomes, B cell receptor VH/VL pairings and DNA oligos associated with antigen tetramers bound by B cells using the 10X Genomics platform. I will analyze flow cytometry data using FlowJo and sequencing data using CellRanger and receptor repertoire analysis pipeline from our lab. I will perform statistical analyses, including principal component analyses, in R Studio. These experiments will be completed by the end of December 2023 and data analyzed by February 2024.
