MHC Array T Cell Assay System for Monitoring Immune Status in Type 1 Diabetes

项目摘要和摘要： 1型糖尿病(T1D)在美国影响100多万人，通常被诊断为 并可能导致失明、心脏病和肾衰竭。T1D干预的一个主要焦点是 自身反应性T细胞的检测和特征，它在胰岛素攻击中起核心作用 产生胰岛细胞。为了使对T1D的医疗干预最有效，应该检测到这种疾病并 在症状出现之前进行治疗。目前识别自身反应性T1D特异性T细胞靶点的方法 细胞速度慢，技术要求高，劳动和试剂密集型，需要消耗大量的T细胞来 测试有限数量的目标。样本量在T1D中是一个极其重要的考虑因素，通常是 检测临床样本时的限制因素，特别是在儿童和青少年无法提供 大样本。这项SBIR建议的目标是开发一种自动T细胞分析测试来识别 并对自身反应性T1D抗原特异的CD4+和CD8+T细胞进行功能表征 数千个靶点同时使用仅2毫升的患者样本。拟议的系统基于 在将正在申请专利的MHC-多肽阵列技术与自动流动细胞检测和 分析系统。该项目的创新方法将把MHC-多肽阵列结合在一个独立的 具有能够快速、多路转换的高度并行的发光检测系统的样品盒 使用比当前方法小得多的样本体积来分析T1D抗原特异性T细胞。这个 该系统的主要进步将是能够目视地检测抗原特异性的CD4+和CD8+ 自身反应性T细胞与MHC-肽复合体结合及其与MHC：T细胞受体结合的相关性 从而激活和分泌多种细胞因子效应分子。目标是创建一个T细胞分析 该系统提供高通量筛选、多参数表征功能，并结合 最先进的集成样品处理，使用方便。目前，还没有可用的系统 提供功能广泛、使用简单且样本量有限的要求 使用MHC多肽阵列对抗T1D的细胞分析系统。

Project Summary and Abstract: Type 1 Diabetes (T1D) affects more than 1 million people in the United States and is usually diagnosed in children and can lead to blindness, heart disease and kidney failure. A major focus of intervention for T1D is on the detection and characterization of autoreactive T cells, which play a central role in the attack on insulin producing islet cells. For medical intervention for T1D to be most effective, the disease should be detected and treated before the onset of symptoms. Current methods to identify the targets of autoreactive T1D-specific T cells are slow, technically demanding, labor- and reagent-intensive, and consume large numbers of T cells to test limited numbers of targets. Sample size is an extremely important consideration in T1D and is often a limiting factor when testing clinical samples, especially as young children and adolescents are unable to give large samples. The objective of this SBIR proposal is to develop an automated T cell analysis assay to identify and functionally characterize autoreactive T1D antigen-specific CD4+ and CD8+ T cells capable of testing thousands of targets simultaneously using as little as 2ml of patient sample. The proposed system is based upon integrating patent pending MHC-peptide array technology with an automated flow cell detection and analysis system. The innovative approach of this project will combine MHC-peptide arrays in a self-contained sample cartridge with a highly parallel luminescent detection system that is capable of rapid, multiplexed analysis of T1D antigen-specific T cells using dramatically smaller sample volumes than current methods. The major advancements of this system will be the ability to visually detect antigen-specific CD4+ and CD8+ autoreactive T cells binding to MHC-peptide-complexes and to correlate MHC: T cell receptor binding to resulting activation and secretion of multiple cytokine effector molecules. The goal is to create a T cell analysis system that offers high throughput screening, multi-parametric characterization capability and incorporates state-of- the-art integrated sample handling for ease-of-use. At present, there are no systems available that offer the breadth of capabilities, the simplicity of use and limited sample size requirements as the proposed T cell analysis system using MHC-peptide arrays in the fight against T1D.