Identifying antibodies (Abs) that neutralize infectious agents is the first step for developing therapeutics, vaccines, and diagnostic tools for these infectious agents. However, current approaches for identifying neutralizing Abs (nAbs) typically rely on dilution-based assays that are costly, inefficient, and only survey a small subset of the entire repertoire. There are also intrinsic biases in many steps of conventional nAb identification processes. More importantly, conventional assays rely on simple Ab-antigen binding assays, which may not result in identifying the most potent nAbs, as the strongest binder may not be the most potent nAb. Droplet microfluidic systems have the capability to overcome such limitations by conducting complex multi-step assays with high reliability, resolution, and throughput in a pico-liter volume water-in-oil emulsion droplet format. Here, we describe the development of PRESCIENT (Platform for the Rapid Evaluation of antibody SucCess using Integrated microfluidics ENabled Technology), a droplet microfluidic system that can enable high-throughput single-cell resolution identification of nAb repertoires elicited in response to viral infection. We demonstrate PRESCIENT's ability to identify Abs that neutralize a model viral agent, Murine coronavirus (murine hepatitis virus), which causes high mortality rates in experimentally infected mice. In-droplet infection of host cells by the virus was first demonstrated, followed by demonstration of in-droplet neutralization by nAbs produced from a single Ab-producing hybridoma cell. Finally, fluorescence intensity analyses of two populations of hybridoma cell lines (nAb-producing and non-nAb-producing hybridoma cell lines) successfully discriminated between the two populations. The presented strategy and platform have the potential to identify and investigate neutralizing activities against a broad range of potential infectious agents for which nAbs have yet to be discovered, significantly advancing the nAb identification process as well as reinvigorating the field of Ab discovery, characterization, and development.

中文翻译：

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当前：使用集成的微流体技术快速评估抗体成功的平台。

鉴定可中和传染原的抗体（Abs）是开发用于这些传染原的治疗剂，疫苗和诊断工具的第一步。但是，当前用于识别中和抗体（nAb）的方法通常依赖于基于稀释的测定，该测定成本高，效率低，并且仅调查整个库中的一小部分。在常规nAb鉴定过程的许多步骤中也存在固有偏差。更重要的是，常规测定法依赖于简单的Ab抗原结合测定法，由于最强的结合剂可能不是最有效的nAb，因此可能无法鉴定出最有效的nAb。液滴微流体系统具有通过进行具有高可靠性，分辨率，和通量为皮微升油包水型乳液滴形式。在这里，我们描述了PRESCIENT（使用集成的微流体启用技术对抗体SucCess进行快速评估的平台）的发展，这是一种微滴微流体系统，可以实现对病毒感染引起的nAb组成部分的高通量单细胞分辨率鉴定。我们证明了PRESCIENT能够识别可中和模型病毒剂鼠科冠状病毒（鼠科肝炎病毒）的抗体的能力，鼠科冠状病毒可导致实验感染小鼠的高死亡率。首先证明了病毒对宿主细胞的液滴内感染，然后证明了由单个产生Ab的杂交瘤细胞产生的nAb对液滴的中和作用。最后，成功区分了两个群体的杂交瘤细胞系（产生nAb和不产生nAb的杂交瘤细胞系）的荧光强度分析。提出的策略和平台具有识别和调查针对尚未发现nAb的广泛潜在传染原的中和活性的潜力，这大大促进了nAb的鉴定过程，并重振了Ab发现，表征和鉴定领域。发展。