Novel Cellular Microarray Assay for Profiling T-Cell Peptide Antigen Specificities
Novel Cellular Microarray Assay for Profiling T-Cell Peptide Antigen Specificities
复制标题
DOI：
10.1021/pr100447b
复制
发表时间：
2010-11-01
影响因子：
4.4
通讯作者：
Schneck, J. P.
中科院分区：
文献类型：
--
作者：
Yue, C.;Oelke, M.;Schneck, J. P.
我们提出了一种新的细胞微阵列分析方法，使用可溶性多肽负载的HLAA2-Ig二聚体复合体，通过保留二聚体复合体的分子灵活性来优化多肽-人类白细胞抗原结合的亲和力，同时获得相对于同源T细胞受体更高的复合体浓度。通过将分子T细胞受体识别事件从导致在微阵列上捕获抗原特异性细胞的结合相互作用中分离出来，在分析开发方面取得了开创性的进展。这一进展使得能够定量确定不同T细胞群体中的抗原特异性频率，而不需要列举在微阵列上捕获的细胞数量。细胞捕获的特异性、对低抗原特异性频率的敏感性和抗原T细胞特异性的定量是使用具有准备的抗原特异性CTL频率的CD8T细胞群体和从外周血中分离的异种T细胞来建立的。结果表明，对于高通量、基础广泛的低频抗原特异性的定量评估，具有几个优点。该检测方法能够使用细胞微阵列来确定人群内和人群间抗原特异性T细胞反应的稳定性和流量。
We present a novel cellular microarray assay using soluble peptide-loaded HLA A2-Ig dimer complexes that optimizes the avidity of peptide-HLA binding by preserving the molecular flexibility of the dimer complex while attaining much higher concentrations of the complex relative to cognate T-cell receptors. A seminal advance in assay development is made by separating the molecular T-cell receptor recognition event from the binding interactions that lead to antigen-specific cell capture on the microarray. This advance enables the quantitative determination of antigen-specific frequencies in heterogeneous T-cell populations without enumerating the number of cells captured on the microarray. The specificity of cell capture, sensitivity to low antigen-specific frequencies, and quantitation of antigenic T-cell specificities are established using CD8 T-cell populations with prepared antigen-specific CTL frequencies and heterogeneous T cells isolated from peripheral blood. The results demonstrate several advantages for high-throughput broad-based, quantitative assessments of low-frequency antigen specificities. The assay enables the use of cellular microarrays to determine the stability and flux of antigen-specific T-cell responses within and across populations.