DEFINING HEPATITIS C VIRUS-SPECIFIC CTL ACTIVITY IN MAN

定义人类丙型肝炎病毒特异性 CTL 活性

• 批准号: 2517373
• 负责人: K J Kao
• 金额: $ 17.32万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-01 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2517373
• 关键词: cell line; chronic disease /disorder; clinical research; cytotoxic T lymphocyte; cytotoxicity; diagnosis design /evaluation; epidemiology; epitope mapping; hepatitis C; hepatitis C virus; human subject; peptide library; phenotype; tissue /cell culture; transfection; vaccinia virus; virus genetics

DESCRIPTION: This proposal has two major specific aims: 1) To demonstrate the existence HCV-specific liver-infiltrating cytotoxic T lymphocytes in chronic HCV infection. The applicant has already determined the HCV genotype of many patients and will complete this. To detect the presence of HCV-specific CTLs, he will employ a target cell consisting EBV immortalized B-lymphoblastoid cells from each patient with chronic HCV. These cells will be infected with vaccinia virus carrying HCV gene inserts coding for specific candidate regions that are suspected of serving as epitopes in the CTL recognition process. CTLs will be obtained by bulk expansion of lymphocytes harvested from liver biopsies. chromium release assay of target cell lysis will be used to detect cells and quantitate the cytotoxic activity. 2) The second specific aim is to define the actual epitopes involved in the CTL- mediated recognition. CTLs identified in specific aim 1 will be cloned for use in screening small synthetically produced peptides overlapping in regions identified in specific aim 1. Sensitization of target cells and the cytolysis assay will be used.

描述：该提案有两个主要具体目标：1) 证明存在 HCV 特异性肝浸润细胞毒性 T 慢性 HCV 感染中的淋巴细胞。 申请人已经 确定了许多患者的 HCV 基因型并将完成这一工作。 为了检测 HCV 特异性 CTL 的存在，他将使用靶细胞 由每位患者的 EBV 永生化 B 淋巴母细胞组成 患有慢性丙肝病毒。 这些细胞会被痘苗病毒感染 携带编码特定候选区域的 HCV 基因插入片段 怀疑在 CTL 识别过程中充当表位。 CTL 将通过大量扩增从肝脏收获的淋巴细胞来获得 活检。 靶细胞裂解的铬释放测定将用于 检测细胞并定量细胞毒活性。 2）第二个 具体目标是定义 CTL 中涉及的实际表位 介导的识别。 将克隆特定目标 1 中确定的 CTL 用于筛选小的合成产生的重叠肽 特定目标1中确定的区域。靶细胞的敏化 并将使用细胞溶解测定。