A microchip to analyze trafficking leukocytes in Alzheimer’s disease patients

 描述（由申请人提供）：阿尔茨海默病（AD）是最常见的神经退行性疾病之一，可导致有害结果，如进行性记忆缺陷和认知障碍。尽管预计AD的患病率在未来30年内将翻一番，但目前还没有广泛接受的用于早期检测或非侵入性监测AD的分子生物标志物。越来越多的证据表明，免疫反应和脑炎症参与了中枢神经系统（CNS）神经退行性疾病的发病机制。参与脑深部炎症反应的免疫细胞经常进入脑脊液（CSF），即所谓的CNS“循环”系统，这些细胞携带有关脑深部炎症病理的信息。另一方面，虽然血脑屏障（BBB）限制免疫细胞进入CNS，但在病理生理状态下，少量免疫细胞可以穿过CNS参与免疫监视。这些免疫细胞的过度迁移或功能异常有助于神经退行性病变的发展。已经假设CSF中的这些运输白细胞是检测和测量炎性神经退行性疾病的潜在细胞标志物。然而，由于（i）运输白细胞的缺乏（约1个细胞/微升）和（2）细胞高度异质性以及细胞分泌的不同免疫效应功能/蛋白质（高达40），其仍然具有挑战性。IsoPlexis拥有一种原型手持技术，首次提供了在单细胞水平上测量许多（多达45种）这些关键效应蛋白的能力。同时，该装置以其设想的形式需要少得多的细胞输入量（~1000），这代表了优于现有单细胞仪器的主要优点（例如，流式细胞仪）用于分析罕见的运输白细胞的特定应用。它也将比现有的单细胞仪器成本低得多，代表了显着的市场优势。因此，我们计划开发一种完全集成的系统，该系统在同一微型设备上使用纳米粗糙表面和单细胞效应蛋白分析来富集运输白细胞，以真正实现广泛使用运输白细胞作为早期诊断和监测炎症性神经退行性疾病（特别是AD）的生物标志物的机会。为了实现这一目标，我们建议：1。在IsoPlexis微型装置中安装细胞捕获模块，以进行低丰度白细胞的芯片上分离，然后进行高度多重免疫功能分析。2.开发一种集成的载体装置，以执行集成微芯片的可靠操作，用于测量来自CSF的罕见运输白细胞。我们期望开发一种独特的微创方法来定量测量脑深部的炎症状况，以用于使用CSF对AD的早期诊断和治疗监测。这种方法也将对临床前或临床应用产生广泛影响，用于常规筛查或监测炎性神经退行性疾病。

 DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is one of the most common neurodegenerative diseases that lead to detrimental outcomes such as progressive memory deficit and cognitive impairment. Although it is expected that the prevalence of AD will double over the next 30 years, currently no widely-accepted molecular biomarkers for early detection or non-invasive monitoring of AD. There has been increasing evidence that immune responses and brain inflammation are involved in the pathogenesis of neurodegenerative disorders in the central nervous system (CNS). The immune cells participating in the inflammatory response in the deep brain often get into cerebrospinal fluid (CSF), so called the "circulatory" system of CNS, and these cells carry the information about deep brain inflammatory pathology. On the other hand, while the blood-brain barrier (BBB) restricts the entry of immune cells into the CNS, a small number of immune cells can traverse into the CNS during pathophysiological states to participate in immune surveillance. Excessive migration or abnormal functioning of these immune cells contribute to the development of neurodegenerative pathology. It has been hypothesized that these trafficking leukocytes in CSF are potential cell markers to detect and measure inflammatory neurodegenerative disease. However, it remains challenging due to (i) the paucity of trafficking leukocytes (~1 cell/microliter) and (2) the high degree of cellular heterogeneity with diverse immune effector functions/proteins secreted by cells (up to 40). IsoPlexis has a prototype hand-held technology that for the first time provides the ability to measure many (up to 45) of these key effector proteins at the single cell level. At the same time, this device in its envisioned form requires much less amount of cell input (~1000) representing a major advantage over the existing single-cell instruments (e.g., flow cytometer) for the specific application toward the analysis of rare trafficking leukocytes. It will also be far less costly tha existing single-cell instruments, representing a significant market advantage. Thus, we plans to develop a fully integrated system that incorporates the enrichment of trafficking leukocytes using nanorough surfaces and single-cell effector protein analysis on the same microdevice to truly enable the opportunity for wide-spread use of trafficking leukocytes as the biomarker for early stage diagnosis and monitoring of inflammatory neurodegenerative diseases (specifically AD). To reach this goal, we propose: 1. Incorporating a cell capture module in the IsoPlexis microdevice to perform on-chip separation of low abundance leukocytes followed by highly multiplexed immune function analysis. 2. Develop an integrated carrier device to perform reliable operation of the integrated microchip for measuring rare trafficking leukocytes from CSF. We expect to develop a unique and minimally invasive approach to quantitatively measure inflammatory conditions in deep brain for early diagnosis and therapeutic monitoring of AD using CSF. This approach will also have broad impact on preclinical or clinical uses for routine screening or monitoring of inflammatory neurodegenerative diseases.