Novel IgE crosslinking-induced luciferase expression method using human-rat chimeric IgE receptor-carrying mast cells

• Published in: Journal of immunological methods 2024-06, Vol.529, p.113682, Article 113682
• Main Authors: Akiyama, Haruyo, Kurisaka, Chisato, Kumasaka, Kenichi, Nakamura, Ryosuke
• Format: Article
• Language: English
• Subjects: Allergen-specific IgE; Allergy diagnosis; Animals; Cell Line; Genes, Reporter; High-affinity IgE receptor; Humans; Immunoglobulin E - immunology; Luciferase; Luciferases - genetics; Luciferases - metabolism; Mast cells; Mast Cells - immunology; Mast Cells - metabolism; Rats; Receptors, Chimeric Antigen - genetics; Receptors, Chimeric Antigen - immunology; Receptors, Chimeric Antigen - metabolism; Receptors, IgE - genetics; Receptors, IgE - immunology; Receptors, IgE - metabolism; Reproducibility of Results
• ISSN: 0022-1759; 1872-7905; 1872-7905
• DOI: 10.1016/j.jim.2024.113682

The measurement of antigen-specific serum IgE is common in clinical assessments of type I allergies. However, the interaction between antigens and IgE won't invariably trigger mast cell activation. We previously developed the IgE crosslinking-induced luciferase expression (EXiLE) method using the RS-ATL8 mast cell line; however, the method may not be sensitive enough in some cases. In this study, we introduced an NF-AT-regulated luciferase reporter gene into the RBL-2H3 rat mast cell line and expressed a chimeric high-affinity IgE receptor (FcεRI) α chain gene, comprising an extracellular domain from humans and transmembrane/intracellular domains from rats. We generated multiple clones expressing the chimeric receptor. Based on their responsiveness and proliferation, we selected the HuRa-40 clone. This cell line exhibited significantly elevated human α chain expression compared to RS-ATL8 cells, demonstrating a 10-fold enhancement of antigen-specific reactivity. Reproducibility across different batches and operators was excellent. Moreover, we observed a detectable response inhibition by an anti-allergy drugs (omalizumab and cyclosporin A). HuRa-40 cells—which carry the human-rat chimeric IgE receptor—comprise a valuable reporter cell line for the EXiLE method. Their versatility extends to various applications and facilitates high-throughput screening of anti-allergy drugs. Crosslinking of IgE antibodies by multivalent antigens is a key event in allergic reactions. We established a novel cell line (HuRa-40) that expresses human-rat chimeric FcεRIα subunit and luciferase reporter gene. HuRa-40 cells enable highly sensitive measurements of IgE crosslinking-induced luciferase expression (EXiLE), compared with previously used RS-ATL8 cells. This is because HuRa-40 cells express abundant FcεRIα subunits. HuRa-40 cells are also well-suited for high-throughput screening of anti-allergic drugs due to their high reproducibility and wide dynamic range. [Display omitted] •A novel human-rat chimeric FcεRI α chain was designed and expressed in rat mast cells.•The HuRa-40 clone abundantly expressed the chimeric receptor, enabling highly sensitive and reproducible EXiLE responses.•HuRa-40 cells are an ideal tool for high-throughput screening of anti-allergy drugs.