1D-, 2D-Gel Electrophoresis, Immunoblotting, and Enzyme-Linked Immunosorbent Assay (ELISA) for the Study of Food Allergens

Abstract

Protein-based methods have been fundamental for the study of food allergens, not only from a mechanistic and diagnostic point of view, but especially with respect to allergen management and food safety. In this chapter, four individual protocols are suggested, relying on one-dimensional, two-dimensional gel electrophoresis, immunoblotting, and enzyme-linked immunosorbent assay (ELISA). The particularities of the proposed protocols are focused on previous research targeting specific allergenic foods, with cow's milk proteins as case studies. Data on the importance of protein extraction and the use of different animal-raised antibodies and/or sera of food-allergic patients are also critically discussed within method development and optimization. The protocols herein described are successful examples applied to the study of cow's milk allergens in complex matrices, although they can be easily developed and optimized for any food allergen or allergenic food.

Keywords: Allergen characterization; Allergen detection; Animal-raised antibodies; IgE-binding capacity; Immunoreactivity; Protein profile; Sera of food-allergic patients; Western blot.