Recombinant expression of human IgE antibody constructs for analysis of antigenic determinants on dust mite allergens

Defining allergenic epitopes recognized by human IgE antibodies has long been a goal of allergen research which could ultimately lead to innovations in allergy therapeutics. The recent development of human IgE monoclonal antibody (mAb) technology (JCI Insight 2018: 3(20)) means that goal is realistically achievable. Our objective was to express recombinant human IgE mAb for mapping allergenic epitopes on the major dust mite allergens, Derp1 and Derp2. Human IgE mAb technology, involving fusion of B-cells from mite-allergic patients with a human myeloma fusion partner using electrical cytofusion, was used to produce monoclonal cell lines expressing unique IgE antibodies. RNA was extracted and sequenced using rapid amplification of cDNA ends. Recombinant antibodies (Fab or chimeric IgE-IgG1) were expressed, purified and tested by ELISA for binding to Derp1 and Derp2. High levels of purified human IgE mAb directed against Derp1 and Derp2 were produced in CHO cells (∼100 mg/L). Chimeric and Fab versions of two non-overlapping anti-Derp2 IgE mAb, 1B8 and 2F10, were compared. Purified 2F10 Fab was 9X more reactive with Derp2 than 1B8 Fab by ELISA. A chimeric IgE-IgG1 construct, 1C14, was expressed and reacted with Der p 1. Crystals for structural studies were obtained from a preparation of Derp1 with 1C14 IgE-IgG1 complex. Recombinant human IgE mAb expressed in mammalian cells will allow clinically relevant epitopes on allergen molecules to be defined. They also provide critical tools for investigation of biologic mechanisms of IgE responses. Understanding the interaction of human IgE antibodies with allergens will facilitate design of hypoallergens.