Simultaneous antigen receptor repertoire profiling and single-cell transcriptomics in T and B lymphocytes from limited clinical samples

对有限临床样本中的 T 和 B 淋巴细胞进行同步抗原受体库分析和单细胞转录组学分析

• 批准号: 8971431
• 负责人: Steven Edward Bosinger
• 金额: $ 39.96万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-26 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8971431
• 关键词: Antibodies; Antigen Receptors; Antigens; Antiviral Response; Area; B-Lymphocytes; Base Sequence; Benchmarking; Binding; Biological; Biological Assay; Biopsy Specimen; Blood; Bone Marrow; CD8B1 gene; Cells; Clinical; Clinical Trials; Cloning; Collection; Computational algorithm; Data; Development; Failure; Gene Expression Profile; Genes; Genomics; Goals; Human; Immune response; Immune system; Immunoglobulins; Individual; Infection; Influenza vaccination; Light; Link; Measures; Messenger RNA; Methodology; Methods; Monitor; Population; Procedures; Protocols documentation; RNA Sequences; Reading; Retrieval; Sampling; Sequence Analysis; System; T cell response; T-Lymphocyte; Technical Expertise; Techniques; Testing; Tissue-Specific Gene Expression; Translating; Vaccination; Vaccines; Validation; Work; arm; base; clinically relevant; cost; cost effectiveness; high throughput screening; high throughput technology; improved; insight; next generation sequencing; prospective; public health relevance; receptor; rectal; single cell sequencing; success; transcriptome sequencing; transcriptomics; vaccine development; vaccine trial; vaccinology

 DESCRIPTION (provided by applicant): In the vast majority of vaccine trials, the primary readouts are empirical do not collect information about the immunological determinants of a vaccine's success or failure. In recent years, several vaccine studies have combined high-throughput transcriptomic data with measures of antigenicity. These "systems vaccinology" studies have proven invaluable insight into the determinants of efficacy and antigenicity, and have demonstrated the enormous value in mechanistic studies of human vaccination. However, one area that is particularly understudied is the composition of antigen specific receptors that arise during successful and unsuccessful vaccinations. Collection of antigen-specific clonotype information in clinical vaccine studies would provide valuable data that could be used to accelerate vaccine development. The utility of current methodology for repertoire sequencing for immunological studies has been limited due to several factors (i) high-cost and low-throughput of cloning based assays (ii) most high-throughput assays only provide information on a single gene(typically the H-chain of immunoglobulin or ß-chain for TCRs), or (iii) do not link ag-receptor sequences with immunophenotypic or transcriptomic data. In preliminary work, we have developed a protocol that simultaneously queries the transcriptome and paired antigen receptor sequences in B lymphocytes derived from human bone marrow after flu vaccination using next generation sequencing. The goal of this proposal is to complete development of this combined "-Seq" assay for both B and T cells, including establishment of its limitations and benchmark against contemporary repertoire sequencing techniques.

 描述（由申请人提供）：在绝大多数疫苗试验中，主要读数是经验性的，不收集有关疫苗成功或失败的免疫学决定因素的信息。近年来，一些疫苗研究已经将高通量转录组学数据与抗原性措施相结合。这些“系统疫苗学”研究已经证明了对效力和抗原性决定因素的宝贵见解，并证明了人类疫苗接种机制研究的巨大价值。然而，一个特别未充分研究的领域是在成功和不成功的疫苗接种期间出现的抗原特异性受体的组成。在临床疫苗研究中收集抗原特异性克隆型信息将提供可用于加速疫苗开发的有价值的数据。由于以下几个因素，当前用于免疫学研究的谱系测序方法的实用性受到限制：（i）基于克隆的测定的高成本和低通量（ii）大多数高通量测定仅提供有关单个基因（通常是免疫球蛋白的H链或β链）的信息。TCR的链），或（iii）不将ag受体序列与免疫表型或转录组学数据联系起来。在初步工作中，我们已经开发了一个协议，同时查询转录组和配对抗原受体序列的B淋巴细胞来源于人骨髓流感疫苗接种后，使用下一代测序。本提案的目标是完成用于B和T细胞的这种组合“-Seq”测定的开发，包括建立其针对当代库测序技术的限制和基准。