Droplet-based Analysis of Secretion and Sorting (DASS) for characterization and selection of individual immune cells

基于液滴的分泌和分选分析 (DASS),用于个体免疫细胞的表征和选择

基本信息

  • 批准号:
    9040978
  • 负责人:
  • 金额:
    $ 6.8万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2015
  • 资助国家:
    美国
  • 起止时间:
    2015-04-01 至 2017-09-30
  • 项目状态:
    已结题

项目摘要

DESCRIPTION (provided by applicant): Methods to analyze the products secreted from single cells will be of particular value for studies of the immune system. A successful antibody response requires activation of the individual cells that encode useful antibodies. Similarly, a successful Cytotoxic-T-Lymphocyte (CTL) response requires activation of the individual cells that encode appropriate T-cell receptors. It has been difficult to study secretion from single cell because the cell and its secreted products must be maintained in the same compartment. Multi-well plates are not suitable because the output of a single cell is too dilute for measurement in the relatively large well volumes (>2 uL). We have developed water-in-oil droplets as a powerful tool to study molecules secreted from single cells: A single cell in one of these small volume (40 pL) droplets can secrete molecules to measurable concentrations in < one hour. These droplets can be formed, analyzed and selected at rates ~1000 droplets per second, allowing for high-speed analysis. We will develop a platform to measure single-cell secretion of Tumor Necrosis Factor Alpha (TNF-� Interferon Gamma (IFN-?), and Granzyme B, key functional and regulatory molecules of the immune system. A major advantage of our water-in-oil droplet platform, relative to well-based systems, is that living cells of interest can be rapidly identifie and collected. These cells can then be used for further study, or they can be expanded to high cell numbers for use in adoptive cell therapy research. These assays will be generally useful to study immune system activation and regulation. In addition, the assays will enable the isolation of individual T-cells with the ability to kill infected or cancerous cells. This will provide a powrful means to determine the protein sequence of useful T- cell receptors. These sequences can be used for Chimeric T-Cell therapy, or used to create antibody-like molecules to enable identification of their cancer-cell targets, some of which might prove to be useful cancer-cell markers. In a second application, droplet-selected CTLs will be released and expanded, and the resulting expanded population will be greatly enriched for cells with autologous anti-cancer-cell activity. This process, or features of it, might be important in developing more effective active adoptive cell therapy protocols. In Phase II research, we will make these tools available through commercializing prototype microfluidic detection and instruments, and by selling quality-controlled assay reagents.
描述(由申请人提供):分析单细胞分泌产物的方法将对免疫系统的研究具有特殊价值。成功的抗体应答需要激活编码有用抗体的单个细胞。同样,成功的细胞毒性t淋巴细胞(CTL)反应需要激活编码适当t细胞受体的单个细胞。由于细胞及其分泌产物必须保持在同一个隔室中,因此研究单细胞的分泌一直是困难的。多孔板不适合,因为单个电池的输出太稀,无法在相对较大的孔体积(bbb20 uL)中进行测量。我们已经开发出油包水液滴作为研究单细胞分泌分子的有力工具:单个细胞在这些小体积(40 pL)液滴中的一个可以在不到一小时内分泌可测量浓度的分子。这些液滴可以以每秒1000个液滴的速度形成、分析和选择,从而实现高速分析。我们将开发一个测量单细胞分泌肿瘤坏死因子α (TNF-干扰素γ (IFN- γ))和颗粒酶B的平台,这些分子是免疫系统的关键功能和调节分子。相对于基于油井的系统,我们的油包水液滴平台的一个主要优势是可以快速识别和收集感兴趣的活细胞。然后这些细胞可以用于进一步的研究,或者它们可以扩展到高细胞数量用于过继细胞治疗研究。这些检测对于研究免疫系统的激活和调节通常是有用的。此外,这些检测将使分离出具有杀死感染细胞或癌细胞能力的单个t细胞成为可能。这将为确定有用T细胞受体的蛋白序列提供一种强有力的手段。这些序列可用于嵌合t细胞治疗,或用于制造抗体样分子以识别其癌细胞目标,其中一些可能被证明是有用的癌细胞标记物。在第二种应用中,液滴选择的ctl将被释放和扩增,扩增后的细胞群将大大丰富,具有自体抗癌活性的细胞。这一过程或其特征可能对开发更有效的主动过继细胞治疗方案很重要。在第二阶段的研究中,我们将通过商业化原型微流体检测和仪器,以及销售质量控制的分析试剂,使这些工具可用。

项目成果

期刊论文数量(1)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Droplet encapsulation improves accuracy of immune cell cytokine capture assays
液滴封装提高了免疫细胞细胞因子捕获测定的准确性
  • DOI:
    10.1039/c9lc01261c
  • 发表时间:
    2020
  • 期刊:
  • 影响因子:
    6.1
  • 作者:
    Yuan, Yuan;Brouchon, Julie;Calvo-Calle, J. Mauricio;Xia, Jing;Sun, Li;Zhang, Xu;Clayton, Kiera L.;Ye, Fangfu;Weitz, David A.;Heyman, John A.
  • 通讯作者:
    Heyman, John A.
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JOHN A HEYMAN其他文献

JOHN A HEYMAN的其他文献

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{{ truncateString('JOHN A HEYMAN', 18)}}的其他基金

Droplet-based Analysis of Secretion and Sorting (DASS) for characterization and selection of individual immune cells
基于液滴的分泌和分选分析 (DASS),用于个体免疫细胞的表征和选择
  • 批准号:
    8832841
  • 财政年份:
    2015
  • 资助金额:
    $ 6.8万
  • 项目类别:
Droplet Compartmentalized Selection for Deep-Mining of Antibody Diversity
抗体多样性深度挖掘的液滴区室化选择
  • 批准号:
    7672772
  • 财政年份:
    2009
  • 资助金额:
    $ 6.8万
  • 项目类别:
CLONING AND EXPRESSION OF FULL LENGTH HUMAN ORFS
全长人类 ORFS 的克隆和表达
  • 批准号:
    6041754
  • 财政年份:
    1998
  • 资助金额:
    $ 6.8万
  • 项目类别:
CLONING AND EXPRESSION OF FULL LENGTH HUMAN ORFS
全长人类 ORFS 的克隆和表达
  • 批准号:
    6210224
  • 财政年份:
    1998
  • 资助金额:
    $ 6.8万
  • 项目类别:

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