Immune Phenotyping Core

摘要 免疫表型核心(核心C)的目标是为不同的项目提供工具和试剂 这将被用来描述疫苗接种和感染的免疫反应。 冠状病毒、流感病毒和登革热病毒。核心将利用现有的最先进的技术 在克拉默实验室建立的血清学技术，以及细胞因子和 外周血单核细胞和人扁桃体的趋化因子血浆图谱和Cytek Aurora光谱流式细胞术图谱分析 组织培养(HC)目前在费尔南德斯-塞斯马实验室使用和优化。以下目标为： 提议：目标1：冠状病毒、流感病毒和登革热抗体反应的特征 病毒接种和感染。核心将提供检测、试剂和协议来测量结合 项目1的SARS-CoV-2和项目2的流感病毒的功能性抗体反应。 此外，对人扁桃体组织培养上清液中IgM、Ig G和Ig A的分泌也有一定的影响 随着SARS-CoV-2疫苗类型的不同(项目1)，流感病毒疫苗和病毒(项目2)将 被评估。目的2：冠状病毒、流感病毒对细胞因子/趋化因子反应的特征 以及登革热病毒疫苗接种和感染。核心将分析细胞因子和趋化因子的水平 在接种/感染人的血浆和人扁桃体组织培养上清液中， 项目1、2和3的不同疫苗。目标3：冠状病毒的细胞反应的特征， 流感病毒和登革热病毒疫苗接种和感染。血吸虫病患者细胞免疫功能的分析 使用光谱流式细胞术，检测接种疫苗/感染者的PBMC随时间的变化。人扁桃体HC会 也可以通过Cytek Aurora频谱流式细胞术进行分析，以捕获早期免疫特征和 丙种球蛋白治疗后与获得性免疫反应相对应的细胞群变化 不同的疫苗。我们将在单细胞水平上获得高分辨率数据，以解决最具挑战性的 细胞群体，包括表达病毒抗原的细胞。PBMC也将受到补充 由基因组核心进行的RNAseq转录组学分析。这些工具将用于生成 代表对疫苗接种和/或感染的纵向免疫反应的免疫特征 研究参与者参加了与数据协调的观察性非干预性队列研究 管理和分析核心(核心E)。使用这些免疫学技术获得的数据将是 由数据管理和分析核心进行分析，将它们跨以下不同系统进行比较 这些项目以及与基因组学核心(核心D)中获得的基因组数据相结合 同样的样本。VIVA项目和核心产生的所有数据，包括免疫表型 核心，将由数据分析核心存入ImmPort。

SUMMARY The goal of the Immune Phenotyping Core (Core C) is to provide tools and reagents to the different projects that will be used to characterize the immune responses induced by vaccinations and infections with coronaviruses, influenza viruses and dengue viruses. The Core will leverage existing state-of-the-art serological techniques established in the Krammer laboratory, as well as multiplex analysis of cytokine and chemokine plasma profile and Cytek Aurora Spectral Flow Cytometry profiling of PBMCs and human tonsillar histocultures (HC) currently used and optimized in the Fernandez-Sesma laboratory. The following aims re proposed: Aim 1: Characterization of antibody responses to coronavirus, influenza virus and dengue virus vaccination and infection. The Core will provide assays, reagents and protocols to measure binding and functional antibody responses against SARS-CoV-2 for Project 1 and influenza viruses for Project 2. Additionally, the secretion of IgM, IgG and IgA in the supernatant of human tonsil histocultures (HC) treated with the different SARS-CoV-2 vaccines types (Project 1), influenza virus vaccines and viruses (Project 2) will be assessed. Aim 2: Characterization of cytokine/chemokine responses to coronavirus, influenza virus and dengue virus vaccination and infection. The Core will analyze the levels of cytokines and chemokines in the plasma of vaccinated/infected individuals and the supernatant of human tonsil histocultures treated with different vaccines for Projects 1, 2 and 3. Aim 3: Characterization of cellular responses to coronavirus, influenza virus and dengue virus vaccination and infection. Analysis of the cellular immune profiles of PBMCs from vaccinated/infected individuals over time, using Spectral Flow cytometry. Human tonsillar HC will be also analyzed by Cytek Aurora Spectral Flow Cytometry in order to capture early immune signatures and changes in cell populations corresponding to adaptive immune responses in those HC after treatment with different vaccines. We will obtain high-resolution data at the single-cell level to resolve the most challenging cell populations including cells expressing viral antigens. PBMCs will also be subjected to a complementary transcriptomics analysis by RNAseq conducted by the Genomics Core. These tools will serve to generate immune signatures representative of the longitudinal immune responses to vaccination and/or infection in study participants enrolled in observational non-interventional cohort studies in coordination with the Data Management and Analysis Core (Core E). Data obtained using these immunological techniques will be analyzed by the Data management and Analysis Core comparing them across the different systems used in the projects as well as in combination with the genomic data obtained in the Genomics Core (Core D) from the same samples. All data generated by the VIVA Projects and Cores, including the Immune Phenotyping Core, will be deposited by the Data Analysis Core into ImmPort.