Background

Allergic asthma is a complex inflammatory disease, the development of which involves multiple intricate gene-environment interactions. A defining factor of the genetic component for disease inception is the predisposition to developing allergen-specific immunoglobulin (Ig) E to common allergens, otherwise known as atopy (Sly et al., 2008). As such, allergic (atopic) sensitization to innocuous perennial aeroallergens during early childhood is now recognized as a fundamental risk factor in the establishment of potentially life-long allergic asthmatic disease (Illi et al., 2006; Sly et al., 2008), particularly when sensitization occurs to multiple aeroallergens (Stoltz et al., 2013).

Murine models have long provided a means to investigate the cellular and molecular mechanisms that drive onset and progression of allergic airway diseases such as allergic asthma (Kumar et al., 2016). In this regard, mice are typically experimentally sensitized to a model antigen, resulting in CD4⁺ T-helper (Th) type 2-driven inflammation and the production of antigen-specific IgE (Mincham et al., 2018); the hallmark feature of allergic sensitization. Quantification of circulating serum antigen-specific IgE from antigen sensitized murine models is routinely performed by enzyme-linked immunosorbent assay (ELISA). However, in our experience, these in vitro assays lack the necessary reproducibility for accurate detection of ovalbumin (OVA)-specific serum IgE in both Brown Norway rat and BALB/c mouse models of allergic airway disease. Furthermore, the use of in vitro techniques disregards the functional activity of antigen-specific IgE in a biologically relevant setting.

To combat the disadvantages of in vitro IgE quantification assays, we herein detail a highly reproducible in vivo passive cutaneous anaphylaxis (PCA) assay which enables both the quantification and assessment of biological activity of OVA-specific IgE within serum samples from OVA-sensitized murine models. In this regard, the localized cutaneous anaphylaxis response observed from an OVA-specific IgE positive sample is indicative of mast cell degranulation driven via OVA-induced cross-linking of OVA-specific IgE (Kawakami and Galli, 2002). The resultant release of mast cell granule-derived proinflammatory mediators exemplified by histamine, prostaglandin, leukotriene and tryptase, ultimately stimulates vascular permeability and the subsequent extravasation of the blue detection dye (Bradding and Arthur, 2016). While this protocol has been described for quantifying OVA-specific IgE in murine serum samples, it could be adapted to assess alternative model antigens.