Monitoring peripheral blood leukocyte and immune responses in health and disease

 描述（由申请方提供）：拟定实验将检验假设：使用Dean Flow Fractionation通过白细胞的内在特性分离白细胞将能够分析微升量全血中的先天性和适应性免疫应答。 急性和慢性炎症反应是许多疾病临床表现的组成部分，但免疫功能的常规临床评估仅基于白细胞计数。这通常用全血细胞计数和白细胞分类计数来测量。在当前的临床实践中，关于免疫调节的关键临床决策是基于白细胞和白细胞亚类的计数而做出的，而不评估白细胞功能。此外，外周静脉全血细胞计数的测量使用3-5 mL。根据全血细胞计数和其他常规实验室检查所需的大量血液，患者可能会发生医源性贫血。迫切需要开发能够评估免疫功能并且需要显著更小体积的血液的测定。在初步结果中，我们已经开发出一种惯性微流体方法来选择性地分离基于非标记的细胞类型特异性物理特征的不同细胞靶标。这种新的方法可以从外周静脉血中分离白细胞和红细胞、血小板、白细胞亚类，并通过串联等介电分离来区分活化的中性粒细胞和未活化的细胞。该多功能体外分离系统具有高通量，能够在系统和细胞免疫功能的耦合分析之前对白细胞进行初步分离。为了解决我们的假设，提出了两个具体的目标：1。设计微升规模的预备惯性微流控分离 外周血白细胞; 2.确定系统和细胞免疫功能的微升规模测定。高通量微流体工程的最新进展使该项目成为可能，为复杂免疫监测的新方法铺平了道路。

 DESCRIPTION (provided by applicant): The proposed experiments will test the hypothesis: The separation of leukocytes by their intrinsic properties using Dean Flow Fractionation will enable analyses of innate and adaptive immune responses in microliter quantities of whole blood. Acute and chronic inflammatory responses are integral to the clinical expression of many diseases, yet routine clinical assessment of immune function is based solely on leukocyte count. This is typically measured with a complete blood count and leukocyte differential count. In current clinical practice, critical clinical decisions regarding immune modulation are made based on the count of leukocytes and leukocyte subclasses without assessment of leukocyte function. In addition, the measurement of a peripheral venous complete blood count uses 3-5 mL. Based on the significant blood volume required for the complete blood count and other routine laboratory testing, patients can develop iatrogenic anemia. There is a critical need to develop assays that enable assessment of immune function and require significantly smaller volumes of blood. In preliminary results, we have developed an inertial microfluidic approach to selectively separate diverse cellular targets that is based on non-labeled, cell type specific physical features. From peripheral venous blood, this new approach can separate leukocytes from erythrocytes and platelets, leukocyte subclasses and distinguish activated neutrophils from unactivated cells by tandem iso-dielectric separation. This multi-functional, extracorporeal separation system is high throughput and capable of preparatory separation of leukocytes prior to coupled analyses of systemic and cellular immune function. To address our hypothesis, two specific aims are proposed: 1. Design microliter scale preparatory inertial microfluidic separation of leukocytes in peripheral blood; 2. Determine microliter scale assays of systemic and cell-based immune function. Recent advances in high-throughput microfluidic engineering enable this project, paving the way for a new approach to complex immune monitoring.