Pdot Enabled Cyclic Imaging Cytometry

Pdot 启用循环成像细胞术

• 批准号: 10080645
• 负责人: Jiangbo Yu
• 金额: $ 14.98万
• 依托单位: LAMPROGEN, INC.
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10080645
• 关键词: Acquired Immunodeficiency Syndrome; Address; Air; Antibodies; Area; Biological Markers; Biology; Blood; CD4 Positive T Lymphocytes; Cell Cycle; Cell Lineage; Cell surface; Cells; Characteristics; Color; Cytometry; Detection; Development; Diagnostic; Dyes; Energy Transfer; Exhibits; Family; Flow Cytometry; Fluorescence; Fluorescence-Activated Cell Sorting; Fluorescent Dyes; Fluorescent Probes; Foundations; Hour; Hydrophobicity; Image; Image Cytometry; Immune; Immunooncology; Immunotherapy; Incidence; Label; Lasers; Light; Malignant Neoplasms; Measurement; Measures; Medicine; Messenger RNA; Methodology; Methods; Modernization; Monitor; Motivation; Nucleic Acids; Particle Size; Performance; Periodicity; Phase; Photons; Polymers; Proteins; Quantum Dots; Reagent; Reporting; Research; Scanning; Scheme; Series; Signal Pathway; Signal Transduction; Staining and Labeling; Stains; Surface; Techniques; Time; Tissue imaging; Tissues; Touch sensation; Translating; Walking; Water; Work; base; cancer biomarkers; fluorescence imaging; imaging approach; imaging biomarker; imaging detection; leukemia/lymphoma; multiplexed imaging; nanoparticle; new technology; precision medicine; prognostic; quantum; stem; transcriptome; treatment response; trend; two-photon

Project Summary For modern flow cytometry, high-throughput multiplexing is extremely important because of the great need in analyzing a large number of biomolecules on and in a single cell. This trend is driven by precision medicine and the need to analyze an ever increasing number of immune-cell subtypes (e.g. in immune-oncology). Highly multiplexed cellular analysis using flow cytometry, however, is challenging. To address this limitation of flow cytometry, we propose to develop a Cyclic Imaging Cytometry (CIC) platform that can be developed to offer a simple automated workflow. The concept behind CIC is straightforward, involving the following steps: 1) Rapid labeling of cells with a large color panel of fluorescent probes against different cellular markers; 2) Perform sensitive and rapid fluorescence imaging; 3) Rapid de-staining of the labeled cells; 4) Iterate steps 1-3 to achieve a large number of imaged biomarkers. While the concept of sequential labeling and de-staining has been explored by many research groups over the years, especially in the context of tissue imaging, this approach thus far has not been able to offer the same level of sensitivity and throughput that is routinely provided by flow cytometry. Recently, we and others have developed fluorescent nanoparticles based on semiconducting polymers called Pdots. The motivation of adapting fluorescent semiconducting polymers into nanoparticle labels stems from a number of favorable characteristics, such as large absorptivity, high quantum yield, fast emission rates, and excellent photostability. The resulting Pdots exhibit extraordinarily high fluorescence brightness, a factor of 102 - 104 higher than conventional dyes, and a factor of 10-103 higher than Qdots depending on the particle size. With the development and availability of Pdots, we believe we can develop CIC to exceed the level of performance offered by flow cytometry. Specifically, the high brightness of Pdots enables high sensitivity imaging and detection of cellular biomarkers, even those present at very low expression levels, and the amplified energy transfer present in Pdots allows the development of a large color panel of Pdots that are both bright with narrow emissions and which can be de-stained efficiently for use in CIC.

项目概要 对于现代流式细胞术，高通量多重分析极其重要，因为 分析单个细胞上和细胞内的大量生物分子的巨大需求。这个趋势是 由精准医学和分析数量不断增加的免疫细胞的需求驱动 亚型（例如免疫肿瘤学）。使用流式细胞术进行高度多重细胞分析， 然而，这是具有挑战性的。为了解决流式细胞术的这一局限性，我们建议开发一种 循环成像细胞计数 (CIC) 平台可开发为提供简单的自动化 工作流程。 CIC 背后的概念很简单，涉及以下步骤： 1) 快速标记 具有针对不同细胞标记的大量荧光探针彩色面板的细胞； 2) 执行 灵敏、快速的荧光成像； 3）标记细胞快速脱色； 4）迭代 步骤 1-3 实现大量生物标记物成像。虽然顺序的概念 多年来，许多研究小组一直在探索标记和脱色，特别是 在组织成像的背景下，这种方法迄今为止还无法提供相同水平的 流式细胞术常规提供的灵敏度和通量。 最近，我们和其他人开发了基于 称为 Pdot 的半导体聚合物。采用荧光半导体的动机 将聚合物制成纳米颗粒标签源于许多有利的特性，例如 吸收率大、量子产率高、发射速率快、光稳定性好。这 所得 Pdot 表现出极高的荧光亮度，高出 102 - 104 倍 比传统染料高出 10-103 倍，具体取决于颗粒尺寸。 随着 Pdot 的开发和可用性，我们相信我们可以开发 CIC 来超越 流式细胞术提供的性能水平。 具体来说，Pdot 的高亮度可以实现高灵敏度成像和检测 细胞生物标志物，即使是那些表达水平非常低的生物标志物，以及放大的能量 Pdot 中存在的转移允许开发 Pdot 的大型彩色面板，这些面板既 明亮，发射范围窄，可有效脱色以用于 CIC。