Core A: Automated Cell/Biochemical Assays Core

通过诱导针对HIV-1的有效和广谱中和抗体的能力 疫苗接种是我们这个时代生物医学研究的主要挑战之一。建议数 HIVRAD项目旨在应对这一挑战。我们的方法基于这样一个概念： 与已知成熟前体的推测生殖系抗体(IGL)结合的免疫原 BNAbs可以启动一种抗体反应，该反应可以被引导使用一系列 顺序免疫原。因此，这笔赠款的重点是评估免疫方案。 在动物模型(项目1)中使用由基于结构的设计和文库选择的免疫原 筛选(项目2)。此项目成功的关键要求包括能够 评估数千份来自免疫动物的血清样本，以确定它们是否 产生特定的和富有成效的反应，产生和筛选数以千计的潜力 免疫原和抗体，以及评估结合和中和效力和广度。在这里我们 描述致力于使用液体处理机器人进行自动表达的科学核心 少量的多种不同的蛋白质和高通量的自动化分析。这个 细胞/生化分析自动化核心将设计和实施基于细胞的自动化和 生化分析以高通量表达蛋白质，表征结合亲和力和 特异性，并评估HIV中和抗体的效力。核心A级人员将 验证、改进和排查自动化蛋白质表达、结合和故障排除协议 用于评估抗体和环境蛋白的体外中和试验 努森茨韦格博士和比约克曼博士。核心将培训和帮助学生、博士后研究员、 研究助理和调查人员分析和解释来自自动化的数据 化验。自动化细胞/生化分析核心将维护两个定制设备的Evo 自由液体处理站和具有高通量能力的表面等离子体共振仪，用于本项目中提出的自动化分析。

The ability to elicit potent and broadly neutralizing antibodies (bNAbs) against HIV-1 through vaccination is one of the prime challenges of biomedical research in our time. The proposed HIVRAD project aims to address this challenge. Our approach is based on the concept that immunogens that bind to inferred germline Abs (iGLs) that are precursors to known mature bNAbs can initiate an Ab response that can be guided to produce bNAbs using a series of sequential immunogens. Therefore the focus of this grant is to evaluate immunization regimens in animal models (Project 1) using immunogens selected by structure-based design and library screening (Project 2). Key requirements for the success of this project include the ability to evaluate thousands of serum samples from immunized animals to determine whether they are producing specific and productive responses, producing and screening thousands of potential immunogens and Abs, and assessing binding and neutralization potency and breadth. Here we describe a scientific core dedicated to using liquid handling robots for automated expression of small quantities of many different proteins and high-throughput automated assays. The Cell/Biochemical Assay Automation Core will design and carry out automated cell-based and biochemical assays to express proteins in high-throughput, characterize the binding affinity and specificity, and evaluate the potency of HIV-neutralizing antibodies. Core A personnel will validate, refine, and trouble-shoot protocols for the automated protein expression, binding, and in vitro neutralization assays and perform assays for evaluating antibodies and Env proteins for Drs. Nussenzweig and Bjorkman. The core will train and assist students, postdoctoral fellows, research assistants, and investigators in the analysis and interpretation of data from automated assays. The Automated Cell/Biochemical Assays Core will maintain two custom-equipped Evo Freedom Liquid handling stations and a surface plasmon resonance instrument with high-throughput capabilities for the automated assays proposed in this project.