DEFINING HEPATITIS C VIRUS-SPECIFIC CTL ACTIVITY IN MAN

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细胞 慢性HCV感染的淋巴细胞。 申请人已 确定了许多患者的HCV基因型，并将完成这项工作。 为了检测HCV特异性CTL的存在，他将使用靶细胞 由来自每个患者的EBV永生化B淋巴母细胞组成 慢性HCV 这些细胞会被牛痘病毒感染 携带编码特定候选区域的HCV基因插入物， 怀疑在CTL识别过程中充当表位。 CTLs 将通过大量扩增从肝脏收获的淋巴细胞获得 活组织检查 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 具体的目的是定义参与CTL的实际表位。 介导的识别。 将克隆特异性目的1中鉴定的CTL 用于筛选合成产生的小肽重叠 在具体目标1确定的区域。 靶细胞致敏 并将使用细胞溶解测定。

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.