Efficient identification of HLA-A*2402–restricted cytomegalovirus-specific CD8 1 T-cell epitopes by a computer algorithm and an enzyme-linked immunospot assay
Efficient identification of HLA-A*2402–restricted cytomegalovirus-specific CD8 1 T-cell epitopes by a computer algorithm and an enzyme-linked immunospot assay
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发表时间：
2001
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通讯作者：
K. Kuzushima;Naomi Hayashi;H. Kimura;T. Tsurumi
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作者：
K. Kuzushima;Naomi Hayashi;H. Kimura;T. Tsurumi
由病毒特异性fic细胞毒性T淋巴细胞(CTL)识别的抗原肽是研究细胞毒性T淋巴细胞反应的有用工具，仅在那些拥有呈现这些抗原肽的主要组织相容性复合体I类分子的人中进行。为了扩大应用范围，一种有效的fi策略在给定的MHC背景下确定这些表位是非常必要的。提出了一种快速而有效的fi策略，通过计算机辅助算法、MHC稳定化和酶联免疫斑点分析组成的多重筛选，确定给定的MHC分子中的CTL表位。从83个候选多肽中鉴定出一个主要的巨细胞病毒特异性ficCTL表位QYDPVAALF，该表位位于HLAA2402分子呈递的其下基质65kd磷酸蛋白(Pp65)的氨基酸序列中。从83个候选多肽中fi。结果表明，在fi-A*2402背景下，巨细胞病毒特异性CTL应答高度集中于pp65。以表达pp65基因或巨细胞病毒的重组痘苗病毒感染的自体fi成纤维细胞为抗原提呈细胞，以fic CD81T细胞为效应物，进行内源性加工和提呈。细胞内干扰素-g产生的流式细胞仪分析显示，HLAA241型和巨细胞病毒阳性供者外周血中有0.04%至0.27%的CD81T细胞是fic的特异性多肽。来自血清阳性供者的外周血中有0.79%的CD81T细胞与反应性T细胞克隆特异结合fi。该多肽可作为研究人类白细胞抗原A*2402阳性人群对CMV的CTL反应的有用试剂。2001年；98：1872-1881)
Antigenic peptides recognized by virus-specific cytotoxic T lymphocytes (CTLs) are useful tools for studying the CTL responses exclusively among those who own the major histocompatibility complex (MHC) class I molecules that present the peptides. For widening the application, an efficient strategy to determine such epitopes in the context of a given MHC is highly desirable. A rapid and efficient strategy is presented for the determination of CTL epitopes in the context of given MHC molecules of interest through multiple screenings consisting of a computer-assisted algorithm and MHC stabilization and enzyme-linked immunospot assays. A major cytomegalovirus (CMV)–specific CTL epitope, QYDPVAALF, in the amino acid sequence of its lower matrix 65 kd phosphoprotein (pp65) presented by HLA-A*2402 molecules was identified from 83 candidate peptides. The results indicate that the CMV-specific CTL response is highly focused to pp65 in the context of HLA-A*2402. Endogenous processing and presentation was confirmed using a peptide-specific CD8 1 T-cell clone as the effectors and autologous fibroblast cells infected with recombinant vaccinia virus expressing pp65 gene or CMV as antigen-presenting cells. Flow cytometric analysis of intracellular interferon-g production revealed 0.04% to 0.27% of CD8 1 T cells in peripheral blood of HLA-A24 1 and CMV-seropositive donors to be specific for the peptide.ThetetramericMHC-peptidecom-plexes specifically bound to the reactive T-cell clone and 0.79% of CD8 1 T cells in peripheral blood from a seropositive donor. The peptide could be a useful reagent to study CTL responses to CMV among populationspositiveforHLA-A*2402.(Blood. 2001;98:1872-1881)