Fc-gamma receptor (FcγR) activation by soluble IgG immune complexes (sICs) represents a major mechanism of inflammation in certain autoimmune diseases such as systemic lupus erythematosus (SLE). A robust and scalable test system allowing for the detection and quantification of sIC bioactivity is missing. Previously described FcγR interaction assays are limited to certain FcγRs, lack scalability and flexibility, are not indicative of receptor activation or lack sensitivity towards sIC size. We developed a comprehensive reporter cell panel detecting individual activation of FcγRs from humans and the mouse. The reporter cell lines were integrated into an assay format that provides flexible read-outs enabling the quantification of sIC reactivity via ELISA or a fast detection using flow cytometry. This identified FcγRIIA(H) and FcγRIIIA as the most sIC-sensitive FcγRs in our test system. Applying the assay we demonstrate that sICs versus immobilized ICs are fundamentally different FcγR-ligands with regard to FcγR preference and signal strength. Reaching a detection limit in the very low nanomolar range, the assay proved also to be sensitive to sIC stoichiometry and size enabling for the first time a complete reproduction of the Heidelberger-Kendall precipitation curve in terms of immune receptor activation. Analyzing sera from SLE patients and mouse models of lupus and arthritis proved that sIC-dependent FcγR activation has predictive capabilities regarding severity of SLE disease. The new methodology provides a sensitive, scalable and comprehensive tool to evaluate the size, amount and bioactivity of sICs in all settings.

中文翻译：

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通过全面的 FcγR 报告细胞面板检测和功能解析可溶性多聚体免疫复合物

可溶性 IgG 免疫复合物 (sIC) 激活 Fc-γ 受体 (FcγR) 是某些自身免疫性疾病如系统性红斑狼疮 (SLE) 炎症的主要机制。缺少允许检测和量化 sIC 生物活性的强大且可扩展的测试系统。先前描述的 FcγR 相互作用分析仅限于某些 FcγR，缺乏可扩展性和灵活性，不指示受体激活或缺乏对 sIC 大小的敏感性。我们开发了一个全面的报告细胞面板，可检测人类和小鼠 FcγR 的个体激活。报告细胞系被整合到一种分析格式中，该格式提供灵活的读数，能够通过 ELISA 或使用流式细胞术进行快速检测来量化 sIC 反应性。这将 FcγRIIA(H) 和 FcγRIIIA 鉴定为我们测试系统中对 sIC 最敏感的 FcγR。应用该检测方法，我们证明 sIC 与固定化 IC 在 FcγR 偏好和信号强度方面是根本不同的 FcγR 配体。达到极低纳摩尔范围内的检测极限，该测定还证明对 sIC 化学计量和尺寸敏感，从而首次完全再现了海德堡-肯德尔沉淀曲线在免疫受体激活方面。分析 SLE 患者和狼疮和关节炎小鼠模型的血清证明，sIC 依赖性 FcγR 激活具有预测 SLE 疾病严重程度的能力。新方法提供了一种灵敏、可扩展和全面的工具来评估所有环境中 sIC 的大小、数量和生物活性。