Validation of Imaging and Electrical Impedance-Based Microfluidic Assays for Cell Sickling

β-珠蛋白基因中的单个突变导致异常镰状珠蛋白（HbS）的产生， 导致镰状细胞病（SCD）。HbS在低O2张力下聚合成刚性纤维， 引起变形的、刚性的红细胞（称为细胞镰状化），并导致异常的血液流变学， 血管闭塞危象和溶血性贫血等。细胞镰状化可作为诊断的生物标志物 SCD和监测生物标志物，以评估化疗和基因治疗的疗效 直接或间接地靶向HbS及其聚合。我们最近开发了一种 基于微流体的缺氧芯片，用于处理少量血液样本（约几μL）， 通过将红细胞置于低O2张力下，在体外快速诱导细胞镰状化（约几分钟）。 使用细胞成像或基于电阻抗的方法， 方法.在概念化项目中，我们将使用加标的 白细胞减少的纯合子镰状细胞样本。在下一阶段，我们将在体外验证 使用纯合和杂合样品的测定，建立相关的操作标准， 自动化数据处理和分析。该项目将产生经验证的体外试验， 快速、无标记测量细胞镰状化，以评估患者对基因治疗的反应 治疗和其它一般靶向HbS聚合的治疗。

A single mutation in the β-globin gene results in production of abnormal sickle globin (HbS), leading to sickle cell disease (SCD). HbS polymerizes into rigid fibers under low O2 tension, causing distorted, rigid red cells (known as cell sickling), and leading to abnormal blood rheology, vasooccclusive crisis, and hemolytic anemia etc. Cell sickling can serve as a diagnostic biomarker of SCD and a monitoring biomarker to assess the efficacy of chemotherapy and gene therapies that target HbS and its polymerization, directly and indirectly. We have recently developed a microfluidics-based hypoxia chip to process small volume of blood specimens (~ few μL) and induce cell sickling in vitro rapidly (~ few min) by subjecting red blood cells to low O2 tension. Assessment of cell sickling is achieved label-free, using cell-imaging or electrical impedance-based methods. In the conceptualization project, we will study assay sensitivity using spiked leukoreduced homozygous sickle cell samples. In the next stage, we will validate both in vitro assays using homozygous and heterozygous samples, establish relevant operating standards, and automate data processing and analysis. The project will result in validated in vitro assays for rapid, label-free measurement of cell sickling to assess patient response to gene-therapy treatments and other treatments targeting HbS polymerization in general.