Microfluidic Immunophenotyping for the Diagnosis of Uveitis and Ocular Cancer

主要研究者：Murthy，Shashi提案编号：0827868本项目的目的是建立一个微流体平台，能够表征玻璃体液的细胞和细胞因子含量。该平台将直接受益于患有葡萄膜炎或原发性眼内淋巴瘤（PIOL）的患者，这些患者需要进行玻璃体液的手术切除（玻璃体活检）以获得这些疾病的详细诊断。玻璃体液的微流体免疫分型是细胞学和流式细胞术的当前方法的有吸引力的替代方案，因为它需要相当少的细胞，并允许通常非常脆弱的细胞的快速分析。 葡萄膜炎和PIOL都是与淋巴细胞侵入眼睛的各个区域相关的疾病。大约230万美国人患有葡萄膜炎，并且在美国，由于葡萄膜炎导致的视力丧失占失明的10-15%。PIOL表现为恶性B淋巴细胞的存在，这可能表明脑、视网膜或中枢神经系统的其他区域中的肿瘤形成。 虽然PIOL是一种罕见的疾病，但由于缺乏早期检测技术，PIOL患者恢复的可能性很低。进行玻璃体活检以区分PIOL和葡萄膜炎，并确定靶向治疗路径。对于PIOL，早期检测允许患者开始化疗和放疗。对于葡萄膜炎，确定其根本原因允许使用特定药物而不是通常会导致严重副作用的广泛靶向免疫抑制剂进行治疗。目前分析玻璃体活检产品的方法只能对15%的患者提供结论性诊断，因为它受到可用于分析的细胞数量少及其脆弱性的限制。为了实现项目目标，将进行以下任务：-设计和制造基于抗体-抗原相互作用的细胞和细胞因子捕获微流体装置。-创建一个微流控免疫分型技术的基础上使用溶解抗体的梯度细胞的趋化偏转。使用模拟从葡萄膜炎或PIOL患者中提取的玻璃体液的细胞和细胞因子含量的模型表征微流体装置的性能。确定每种疾病类型的多种标志物的靶细胞和细胞因子浓度的下限。从患者玻璃体活检样本中获得的玻璃体液中的免疫表型细胞和细胞因子。智力优点：如果成功，拟议的研究将提供一种新的和变革性的方法来快速识别与葡萄膜炎和PIOL相关的细胞和细胞因子，克服当前分析技术的缺点。所提出的微流体方法的一部分是基于技术变革的梯度设计，以识别单个细胞上的多个细胞表面标记，而不需要任何标记。 从非常小的样本量中确定细胞和细胞类型的能力将允许PIOL的早期诊断，这不能使用当前技术进行。PI在微流控细胞分析领域已经工作了5年多，该项目是与波士顿Schepens眼科研究所的眼科医生Kameran Laskhari博士合作完成的。更广泛的影响：（a）如果成功，拟议的研究将为葡萄膜炎或PIOL患者提供准确快速的诊断技术。(b)拟议的研究将涉及高中学生和科学教师通过现有的青年学者和RET计划在东北大学的实验研究，（c）涉及实验研究的本科生。(d)将鼓励参与拟议研究的高中生开发科学博览会项目，并从事生物医学科学和工程方面的职业。(e)通过网站传播研究结果并描述葡萄膜炎和PIOL治疗的意义。

PI: Murthy, ShashiProposal Number: 0827868The objective of this project is to create a microfluidic platform capable of characterizing the cellular and cytokine content of vitreous humor. This platform will be of direct benefit to patients suffering from uveitis or primary intraocular lymphoma (PIOL) who need to undergo surgical removal of the vitreous humor (vitreous biopsy) in order to obtain a detailed diagnosis of these diseases. Microfluidic immunophenotyping of vitreous humor is an attractive alternative to the current approaches of cytology and flow cytometry because it requires considerably fewer cells and allows rapid analysis of cells that are typically very fragile. Both uveitis and PIOL are diseases associated with the invasion of lymphocytes into various regions of the eye. Approximately 2.3 million Americans suffer from uveitis, and vision-loss due to uveitis accounts for 10-15% of blindness in the U.S. PIOL is manifested by the presence of malignant B-lymphocytes which may indicate tumor formation in the brain, retina, or other areas of the central nervous system. Although PIOL is an uncommon disease, patients of PIOL have a low likelihood of recovery because of the lack of early detection techniques.The vitreous biopsy is performed to distinguish PIOL from uveitis and identify a targeted therapeutic path. For PIOL, early detection allows patients to begin chemo- and radio-therapy. For uveitis, identification of its underlying cause allows treatement with specific drugs instead of broad-target immunosuppressive agents that typically cause severe side effects. The current approach to analyze the vitreous biopsy product provides a conclusive diagnosis in only 15% of patients because it is constrained by the small number of cells available for analysis and their fragility.The following tasks will be carried out to accomplish the project objective:- Design and fabricate microfluidic devices for cell and cytokine capture based on antibody-antigen interactions.- Create a microfluidic immunophenotyping technique based on the chemotactic deflection of cells using gradients of dissolved antibodies.- Characterize the performance of the microfluidic devices using a model that mimics the cellular and cytokine content of vitreous humor extracted from patients of uveitis or PIOL.- Determine lower bounds of target cell and cytokine concentration for the multiple markers of each disease type.- Immunophenotype cells and cytokines in vitreous fluid obtained from patient vitreous biopsy samples.Intellectual Merit: If successful, the proposed research will provide a new and transformative method to rapidly identify cells and cytokines associated with uveitis and PIOL, overcoming the shortcomings of current analysis techniques. Part of the proposed microfluidic approach is technologically transformative gradient-based design to identify multiple cell surface markers on individual cells without requiring any labeling. The ability to determine cell- and cytokine-type from very small sample sizes will allow for early diagnosis of PIOL, which cannot be performed using current techniques. The PI has worked in the area of microfluidic cell analysis for over 5 years and this project is a collaborative effort between with Dr. Kameran Laskhari, an ophthalmic surgeon at the Schepens Eye Research Institute in Boston.Broader Impact: (a) If successful, the proposed research will provide an accurate and rapid diagnosis technique for patients with uveitis or PIOL. (b) The proposed research will involve high school students and science teachers in experimental research via existing Young Scholar and RET programs at Northeastern University, and (c) involve undergraduates in experimental research. (d) High school students participating in the proposed research will be encouraged to develop science fair projects and pursue careers in biomedical science and engineering. (e) Disseminate research results and describe implications for uveitis and PIOL therapeutics through a website.