CORE 2: Protein Expression and Purification Core

综述-核心2：蛋白质表达和纯化核心 支持艰难梭状芽胞杆菌疫苗的Vanderbilt抗体和抗原发现(Vandy-CDV) 目标，蛋白质表达和纯化核心(Protein Core)将改进和建立蛋白质 表达和纯化策略，以实现对项目1和项目2的抗原和抗体的传递 这项建议的第二条。总体而言，蛋白质核心将有三个目标。第一个目标是表达和纯化C. 艰难梭菌毒素和其他抗原，用作项目1、目标1和候选疫苗的分选试剂和候选疫苗 2，和项目2，目标1。第二个目标将是表达和纯化项目中确定的抗体 1，目标1，和项目2，目标2。抗体的表达和纯化将在μg和mg标度上进行，允许 纯化了数百个克隆。这将支持抗体和抗原的验证和中和。 在项目1的两个目标中进行实验。第三个目标将支持项目1中的结构生物学工作。 生物学已经成为疫苗开发的重要组成部分。在本应用中，抗原 将利用发现和抗原优化来生产新的候选疫苗，重点是开发 缺乏免疫优势但非中和表位的亚基。这些亚单位疫苗候选者将是 经过优化并用于结构研究。我们将鉴定抗体抗原对并生产分子Fab 项目1，目标1中结构研究的碎片和适当的抗原。

Summary – Core 2: Protein Expression and Purification Core To support the Vanderbilt Antibody and Antigen Discovery for Clostridioides difficile Vaccines (VANDy-CdV) goals, the Protein Expression and Purification Core (Protein Core) will improve and establish protein expression and purification strategies to enable delivery of both antigens and antibodies for both Projects 1 and 2 of this proposal. Overall, the Protein Core will have three goals. The first goal is to express and purify C. difficile toxins and other antigens for use as sorting reagents and vaccine candidates in Projects 1, aim 1 and 2, and Project 2, aim 1. The second goal will be expression and purification of antibodies identified in Projects 1, aim 1, and Project 2, aim 2. Antibody expression and purification will be done on the μg and mg scale allowing purification of hundreds of clones. This will support antibody and antigen validation and neutralization experiments in both aims of Project 1. The third goal will support structural biology efforts in Project 1. Structural Biology has become an essential component of vaccine development. In the present application antigen discovery and antigen optimization will be used to produce new vaccine candidates with a focus on developing subunits that lack immunodominant yet non-neutralizing epitopes. These subunit vaccine candidates will be optimized and used for structural studies. We will identify antibody antigen pairs and produce molecular Fab fragments and appropriate antigens for structural studies in Project 1, aim 1.