The stalk of the influenza virus hemagglutinin (HA) is an attractive target for antibody-based universal influenza virus vaccine development. While antibodies that target this part of the virus can be neutralizing, it has been shown in recent years that Fc receptor-mediated effector functions are of significant importance for the protective effect of anti-stalk antibodies. Several assays to measure Fc-Fc receptor interaction-based effector functions like antibody-dependent cellular cytotoxicity and antibody-dependent cellular phagocytosis exist, but they suffer from limitations such as low throughput and high run-to-run variability. Reporter assays for antibody-dependent cellular cytotoxicity based on reporter cells that express luciferase upon engagement of human FcγRIIIa with the Fc of antigen-bound antibodies have been developed as well. These reporter assays can be used in a higher throughput setting with limited run-to-run assay variability but since they express only one Fc receptor, their biological relevance is unclear. Here we optimized an antibody-dependent cellular cytotoxicity reporter assay to measure the activity of antibodies to the conserved stalk domain of H1 hemagglutinin. The assay was then correlated to a CD107a-based degranulation assay, and a strong and significant correlation could be observed. This data suggests that the FcγRIIIa-based reporter assay is a good substitute for functional assays, especially in settings where larger sample numbers need to be analyzed.

中文翻译：

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在基于流感病毒血凝素茎的ADCC报告基因分析中，人血清抗体的活性与CD107a脱粒分析中的活性相关。

流感病毒血凝素（HA）的茎是基于抗体的通用流感病毒疫苗开发的有吸引力的目标。尽管靶向病毒这一部分的抗体可以被中和，但近年来已证明，Fc受体介导的效应子功能对于抗茎秆抗体的保护作用非常重要。存在几种用于测量基于Fc-Fc受体相互作用的效应子功能的测定法，例如抗体依赖性细胞的细胞毒性和抗体依赖性细胞的吞噬作用，但它们存在诸如低通量和高运行差异性的局限性。也已经开发了基于报告细胞的抗体依赖性细胞毒性的报告测定，所述报告细胞在人FcγRIIIa与抗原结合抗体的Fc结合后表达荧光素酶。这些报道基因测定可在较高的通量设置中使用，且每次测定之间的变异性有限，但由于它们仅表达一种Fc受体，因此其生物学相关性尚不清楚。在这里，我们优化了抗体依赖性细胞毒性报告基因测定法，以测量针对H1血凝素保守域的抗体的活性。然后将该测定与基于CD107a的脱粒测定相关，并且可以观察到强烈而显着的相关性。该数据表明，基于FcγRIIIa的报告基因检测可以很好地替代功能性检测，尤其是在需要分析大量样本的环境中。在这里，我们优化了抗体依赖性细胞毒性报告基因测定法，以测量针对H1血凝素保守域的抗体的活性。然后将该测定与基于CD107a的脱粒测定相关，并且可以观察到强烈而显着的相关性。该数据表明，基于FcγRIIIa的报告基因检测可以很好地替代功能性检测，尤其是在需要分析大量样本的环境中。在这里，我们优化了抗体依赖性细胞毒性报告基因测定法，以测量针对H1血凝素保守域的抗体的活性。然后将该测定与基于CD107a的脱粒测定相关，并且可以观察到强烈而显着的相关性。该数据表明，基于FcγRIIIa的报告基因检测可以很好地替代功能性检测，尤其是在需要分析大量样本的环境中。