Lumicks C-Trap Optical Tweezers with Confocal Fluorescence Microscope

Lumicks C-Trap 光镊与共焦荧光显微镜

基本信息

项目摘要

Title: Lumicks C-Trap optical tweezers with confocal fluorescence microscope. PI: Bennett Van Houten, PhD Abstract This proposal requests the purchase of a Lumicks C-trap correlative single-molecule and force microscope, which consists of four optical traps, and a 3-color confocal fluorescence platform for Major and Minor user groups at the University of Pittsburgh and Carnegie Mellon University. The C-trap is the only commercially available instrument that fully integrates laser traps with confocal microscopy. Single molecule analysis has helped revolutionize the field of protein-DNA interactions and each of the users have a focused funded project that would be greatly facilitated by access to this instrument. Through a lease agreement we have been using the instrument since Feb of 2021 and several user groups have successfully generated preliminary data on the instrument with up to three different fluorescent proteins. This instrument represents the first placement of this novel technology within the entire Common Wealth of Pennsylvania, and the new four laser trap module is planned to be added June 2022 to keep up with this cutting-edge technology. The C-trap allows investigators to use biophysical approaches to interrogate protein-DNA interactions and follow molecular motors and assembly of protein machines at specific sites on DNA. This instrument combines several unique features that makes single molecule and force measurements accessible to wide group of users that might be relatively new to the single molecule field. The instrument has an automated microfluidic system with a five chamber imaging flow cell, which provides a wide variety of experimental designs. The instrument is fully temperature controlled so kinetics of protein-DNA interactions can be followed at different temperatures. The laser traps have high temporal and spatial control and can measure down to sub pN forces accurately. The force measurements are real-time correlated with the confocal scanning which can scan at speeds up to 5 msec per scan allowing to capture rapid single molecule kinetics. The software that controls the instrument and allows data analysis is easy to use through a Python user interface. Scripting of experimental work-flow as well as data analysis allows highly reproducible conditions to be established. The two user groups consists of members of the Genome Stability and Cancer Virology Programs at the UPMC-Hillman Cancer Center, where the instrument is housed, as well as other minor users throughout the campus. These groups are interested in the role of DNA damage and repair in cancer and neurodegenerative diseases, as well as how specific viruses can cause cancer. Five Major users have been identified for the purpose of this application and they in turn have requested a majority of the time. The five Minor users will work on the instrument in the remaining time. The Lumicks C-trap is being managed as a user facility at the Hillman Cancer Center and will afford the capacity for advanced single molecule and mechanical force measurements and will ensure sustained and efficient use by faculty working in the biomedical sciences in Pittsburgh
标题:Lumicks C-Trap光学镊子与共聚焦荧光显微镜。 PI:班尼特货车Houten,博士 摘要 本提案要求采购Lumicks C-trap相关单分子和力显微镜, 它由四个光学陷阱和一个三色共聚焦荧光平台组成,供主要和次要用户使用。 匹兹堡大学和卡内基梅隆大学的研究小组。C型捕集器是唯一一个 这是一种将激光陷阱与共聚焦显微镜完全集成的仪器。单分子分析 帮助彻底改变了蛋白质-DNA相互作用领域,每个用户都有一个重点资助的项目, 如果能够利用这一文书,将大大便利这一工作。通过我们一直使用的租赁协议 自2021年2月以来,该仪器和几个用户组已经成功地生成了关于 最多可使用三种不同的荧光蛋白。这个仪器代表了第一次放置这个 新技术在整个共同财富的宾夕法尼亚州,和新的四个激光陷阱模块是 计划于2022年6月加入,以跟上这项尖端技术。C-trap可以让调查人员 使用生物物理学方法来询问蛋白质-DNA相互作用并跟踪分子马达, 在DNA的特定位点组装蛋白质机器。该仪器结合了几个独特的功能, 使单分子和力的测量可以被相对较新的广大用户使用 到单分子领域。该仪器具有带五室成像的自动微流体系统 流动池,它提供了各种各样的实验设计。该仪器完全温度控制 因此可以在不同温度下跟踪蛋白质-DNA相互作用的动力学。激光陷阱具有高 并且可以精确地测量到亚pN力。力测量值为 与共焦扫描实时相关,共焦扫描可以以每次扫描高达5毫秒的速度扫描, 捕获快速单分子动力学。控制仪器并进行数据分析的软件是 易于通过Python用户界面使用。编写实验工作流程和数据分析的脚本 允许建立高度可再现的条件。这两个用户组由 UPMC-Hillman癌症中心的基因组稳定性和癌症病毒学项目,该仪器 以及整个校园的其他小用户。这些组织对DNA的作用感兴趣 癌症和神经退行性疾病的损伤和修复,以及特定病毒如何引起 癌本申请已确定了五个主要用户,他们依次 要求大部分时间。五个次要用户将在剩余时间内使用仪器。的 Lumicks C-trap作为希尔曼癌症中心的用户设施进行管理, 先进的单分子和机械力测量,并将确保持续和有效地使用, 在匹兹堡从事生物医学科学的教师

项目成果

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Bennett Van Houten其他文献

Bennett Van Houten的其他文献

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{{ truncateString('Bennett Van Houten', 18)}}的其他基金

Watching cooperative interactions between base and nucleotide excision repair proteins
观察碱基和核苷酸切除修复蛋白之间的协同相互作用
  • 批准号:
    10355532
  • 财政年份:
    2020
  • 资助金额:
    $ 55.23万
  • 项目类别:
Watching cooperative interactions between base and nucleotide excision repair proteins
观察碱基和核苷酸切除修复蛋白之间的协同相互作用
  • 批准号:
    10171852
  • 财政年份:
    2020
  • 资助金额:
    $ 55.23万
  • 项目类别:
Watching cooperative interactions between base and nucleotide excision repair proteins
观察碱基和核苷酸切除修复蛋白之间的协同相互作用
  • 批准号:
    10582554
  • 财政年份:
    2020
  • 资助金额:
    $ 55.23万
  • 项目类别:
Watching cooperative interactions between base and nucleotide excision repair proteins
观察碱基和核苷酸切除修复蛋白之间的协同相互作用
  • 批准号:
    10763255
  • 财政年份:
    2020
  • 资助金额:
    $ 55.23万
  • 项目类别:
Intersection of NER and DNA crosslink repair processes
NER 和 DNA 交联修复过程的交叉点
  • 批准号:
    8528940
  • 财政年份:
    2013
  • 资助金额:
    $ 55.23万
  • 项目类别:
DNA damage recognition by nucleotide excision repair proteins
核苷酸切除修复蛋白识别 DNA 损伤
  • 批准号:
    8204744
  • 财政年份:
    2010
  • 资助金额:
    $ 55.23万
  • 项目类别:
DNA damage recognition by nucleotide excision repair proteins
核苷酸切除修复蛋白识别 DNA 损伤
  • 批准号:
    8391231
  • 财政年份:
    2010
  • 资助金额:
    $ 55.23万
  • 项目类别:
DNA damage recognition by nucleotide excision repair proteins
核苷酸切除修复蛋白识别 DNA 损伤
  • 批准号:
    8021532
  • 财政年份:
    2010
  • 资助金额:
    $ 55.23万
  • 项目类别:
DNA damage recognition by nucleotide excision repair proteins
核苷酸切除修复蛋白识别 DNA 损伤
  • 批准号:
    8897805
  • 财政年份:
    2010
  • 资助金额:
    $ 55.23万
  • 项目类别:
DNA damage recognition by nucleotide excision repair proteins
核苷酸切除修复蛋白识别 DNA 损伤
  • 批准号:
    8580937
  • 财政年份:
    2010
  • 资助金额:
    $ 55.23万
  • 项目类别:

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