Phage Immunoprecipitation Sequencing (PhIP-Seq) for Analyzing Antibody Epitope Repertoires Against Food Antigens

Abstract

While thousands of food and environmental allergens have been reported, conventional methods for allergy testing typically rely on measuring immunoglobulin E (IgE) binding against panels of dozens to hundreds of antigens. Beyond IgE, also the specificity of other Ig (sub-)classes such as IgG4, has gained interest because of a potential protective role toward allergy. Phage immunoprecipitation sequencing (PhIP-Seq) allows to study hundreds of thousands of rationally selected peptide antigens and to resolve binding specificities of different Ig classes. This technology combines synthetic DNA libraries encoding antigens, with the display on the surface of T7 bacteriophages and next-generation sequencing (NGS) for quantitative readouts. Thereby binding of entire Ig repertoires can be measured to detect the exact epitopes of food allergens and to study potential cross-reactivity. In this chapter, we provide a summary of both the key experimental steps and various strategies for analyzing PhIP-Seq datasets, as well as comparing the advantages and disadvantages of this methodology for measuring antibody responses against food antigens.