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

细胞镰化成像和基于电阻抗的微流控分析的验证

• 批准号: 10001658
• 负责人: E Du
• 金额: $ 4.37万
• 依托单位: FLORIDA ATLANTIC UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-20 至 2020-02-20
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10001658
• 关键词: Automatic Data Processing; Biological Assay; Biological Markers; Blood Volume; Blood specimen; Cellular Assay; Data Analyses; Erythrocytes; Fiber; Genes; Globin; Hemolytic Anemia; Hypoxia; Image; In Vitro; Label; Measurement; Methods; Microfluidics; Monitor; Mutation; Polymers; Process; Production; Sampling; Sickle Cell; Sickle Cell Anemia; Testing; Validation; base; beta Globin; blood rheology; cellular imaging; chemotherapy; diagnostic biomarker; electric impedance; gene therapy; in vitro Assay; patient response; polymerization; sickling; targeted treatment

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.

β-珠蛋白基因的单一突变会导致异常镰状珠蛋白 (HbS) 的产生， 导致镰状细胞病 (SCD) 在低氧张力下聚合成刚性纤维， 导致红细胞扭曲、僵硬（称为细胞镰状），并导致血液流变学异常， 血管闭塞危象、溶血性贫血等。镰状细胞可以作为诊断生物标志物 SCD 和监测生物标志物以评估化疗和基因疗法的功效 我们最近开发了一种直接和间接针对 HbS 及其聚合的方法。 基于微流控的缺氧芯片可处理少量血液样本（~几μL）和 通过使红细胞处于低氧张力下，在体外快速（约几分钟）诱导细胞镰状化。 使用细胞成像或基于电阻抗的方法，无需标记即可实现细胞镰状化的评估 在概念化项目中，我们将使用加标研究测定灵敏度。 在下一阶段，我们将在体外验证去白细胞纯合镰状细胞样本。 使用纯合子和杂合子样本进行测定，建立相关操作标准， 并自动化数据处理和分析，该项目将产生经过验证的体外测定。 快速、无标记测量细胞镰状细胞，以评估患者对基因治疗的反应 一般而言，针对 HbS 聚合的治疗和其他治疗。