Division of Pharmacotherapeutics, Faculty of Pharmaceutical Sciences, Teikyo Heisei University, Tokyo, Japan.
Division of Pharmacotherapeutics, Faculty of Pharmaceutical Sciences, Teikyo Heisei University, Tokyo, Japan.
J Immunol Methods. 2024 Jun;529:113682. doi: 10.1016/j.jim.2024.113682. Epub 2024 May 4.
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.
在 I 型过敏的临床评估中，抗原特异性血清 IgE 的测量很常见。然而，抗原和 IgE 之间的相互作用并不一定会引发肥大细胞的激活。我们之前使用 RS-ATL8 肥大细胞系开发了 IgE 交联诱导的荧光素酶表达（EXiLE）方法；然而，在某些情况下，该方法可能不够灵敏。
在这项研究中，我们将 NF-AT 调节的荧光素酶报告基因引入 RBL-2H3 大鼠肥大细胞系，并表达了一种嵌合高亲和力 IgE 受体（FcεRI）α链基因，该基因包含来自人类的细胞外结构域和来自大鼠的跨膜/细胞内结构域。
我们生成了多个表达嵌合受体的克隆。根据它们的反应性和增殖性，我们选择了 HuRa-40 克隆。与 RS-ATL8 细胞相比，该细胞系显著增加了人类α链的表达，表现出抗原特异性反应性提高了 10 倍。不同批次和操作人员之间的重现性非常好。此外，我们观察到抗过敏药物（奥马珠单抗和环孢素 A）可抑制这种反应。
携带人-大鼠嵌合 IgE 受体的 HuRa-40 细胞构成了 EXiLE 方法的一种有价值的报告细胞系。它们的多功能性扩展到各种应用，并促进了抗过敏药物的高通量筛选。
