Flow lysometry and its biomedical applications

流式溶酶仪及其生物医学应用

• 批准号: 7669133
• 负责人: Frans A. Kuypers
• 金额: $ 20万
• 依托单位: CHILDREN'S HOSPITAL & RES CTR AT OAKLAND
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-15 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7669133
• 关键词: ATP phosphohydrolase; Calcium; Cell membrane; Cell surface; Cells; Cellular biology; Characteristics; Complex; Cytolysis; Data; Detection; Development; Device or Instrument Development; Devices; Engineering; Erythrocytes; Evaluation; Event; Flow Cytometry; Goals; Human; Individual; Kinetics; Lasers; Length; Life; Measurement; Measures; Microscopy; Mus; Nanotechnology; Pathology; Phosphatidylserines; Physiologic pulse; Population; Population Heterogeneity; Reaction; Reporting; Research; Science; Sickle Cell; Signal Transduction; Surface; Suspension substance; Suspensions; System; Techniques; Technology; Testing; base; cell type; design; efficacy testing; monolayer; new technology; novel; research study; single cell analysis

DESCRIPTION (provided by applicant): Single-cell analysis by flowcytometry has revolutionized the measurement of complex cell populations. Only limited probes are available for the measurement of cytosolic components in single cells. While cellular components such as ATP can be measured sensitively in a suspension of cells with permeabilized plasma membranes, the determination of the distribution of such compounds in individual cells in a heterogeneous population is not available. We propose the development a novel technique, flow-lysometry, by a multi- disciplinary approach, combining cell biology and nanotechnology developed by micromechanical engineering, to measure cytosolic components of single cells. Our preliminary data show that we can establish lysis of individual cells in a flow channel and detect ATP by a chemiluminescent pulse similar to an event in flowcytometry. The synchronization of conventional flowcytometry with flow-lysometry will allow measurement of surface components in conjunction with cytosolic components in a heterogeneous cell population. As specific aims we propose to 1. develop a device for flow-lysometry, 2. synchronize flow-lysometry with flowcytometry, and 3. test flow-lysometry on a heterogeneous cell population. Fabrication of dedicated micro fluidic channel flow-lysometry chips interfaced with a conventional flowcytometer will allow synchronized cell- detection, cell-lysis, and cytosolic component sensing. We will test the efficacy of this technology by measuring ATP levels in phosphatidylserine (PS) exposing cells in the heterogeneous population of sickle red blood cells. Our research will establish this novel technology to measure cytosolic components in defined, (subpopulations of) individual cells. We envision that this technique can be applied to various types of cells measuring many different cytosolic components.

描述（由申请人提供）： 通过流式细胞术进行的单细胞分析彻底改变了复杂细胞群的测量。只有有限的探针可用于测量单细胞中的胞质组分。虽然细胞组分如ATP可以在具有透性质膜的细胞悬浮液中灵敏地测量，但不能确定这些化合物在异质群体中的单个细胞中的分布。我们建议开发一种新的技术，流动测液法，通过多学科的方法，结合细胞生物学和微机械工程开发的纳米技术，以测量单细胞的胞质组分。我们的初步数据表明，我们可以建立一个流动通道中的单个细胞的裂解和检测ATP的荧光脉冲类似于在流式细胞仪的事件。常规流式细胞术与流式细胞仪的同步将允许测量异质细胞群体中的表面组分与胞质组分。作为具体目标，我们建议1。开发流动测渗装置，2.使流式细胞术与流式细胞术同步，以及3.在异质细胞群上测试流动-溶量测定法。制造与常规流式细胞仪连接的专用微流体通道流动-溶解测定芯片将允许同步的细胞检测、细胞溶解和细胞溶质组分感测。我们将通过测量镰状红细胞异质群体中磷脂酰丝氨酸（PS）暴露细胞中的ATP水平来测试该技术的功效。我们的研究将建立这种新的技术来测量定义的单个细胞（亚群）中的胞质组分。我们设想这种技术可以应用于测量许多不同的胞质组分的各种类型的细胞。

项目成果

Eukaryotic protein recruitment into the Chlamydia inclusion: implications for survival and growth.

真核蛋白招募到衣原体内含物中：对生存和生长的影响。

• DOI: 10.1371/journal.pone.0036843
• 发表时间: 2012
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Soupene,Eric;Rothschild,James;Kuypers,FransA;Dean,Deborah
• 通讯作者: Dean,Deborah