Ultrasound-enabled two-photon FRET microscopy

超声双光子 FRET 显微镜

• 批准号: 8114586
• 负责人: Jerome Mertz
• 金额: $ 8.16万
• 依托单位: BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8114586
• 关键词: Affect; Algorithms; Amplifiers; Animal Model; Caenorhabditis elegans; Cell physiology; Communities; Coupled; Detection; Environment; Fluorescence; Fluorescence Resonance Energy Transfer; Frequencies; Functional Imaging; Generations; Goals; Hemorrhage; Hybrids; Image; Label; Lighting; Measurement; Mechanics; Membrane Potentials; Microbubbles; Microscopy; Molecular; Molecular Conformation; Optics; Photons; Process; Reaction Time; Reporter; Research Technics; Sampling; Signal Transduction; Solutions; Techniques; Testing; Thick; Tissues; Ultrasonic wave; Ultrasonography; base; fluorescence microscope; improved; in vivo; nanoscale; research study; sound; tool; two-photon

DESCRIPTION (provided by applicant): Fluorescence resonance energy transfer (FRET) can serve as a nanoscale molecular ruler. When used in imaging applications, it is a highly sensitive reporter of donor-acceptor molecular configuration. In most cases, FRET is utilized with standard one-photon excitation. Extensions of FRET to two-photon excitation are generally hampered by the problem of donor and acceptor bleed through, which imposes the requirement of technically complicated spectral unmixing algorithms or fluorescence lifetime measurements. We propose to considerably simplify the detection of FRET with two-photon excitation. Our goal is to establish the feasibility of modulating FRET signal using ultrasonic waves. Since FRET is sensitive to distance between donor-acceptor pairs at the nanoscale, mechanical compression and tension produced by sound waves are likely to modulate the FRET signal, serving as a useful signature to distinguish FRET from donor and acceptor bleed-through, which is a particular problem in multiphoton FRET. In particular, we propose to conduct preliminary experiments to establish the premise that ultrasound modulates FRET. We will investigate two types of samples: genetically encoded FRET probes in tissue (or tissue-like environments), and labeled microbubbles in solution. In the former case, we will investigate the application of our technique to in-vivo imaging in C.Elegans. FRET is a powerful tool for functional imaging. We believe that our hybrid ultrasound-optical technique will significantly improve the isolation of FRET signals and therefore be of benefit to any FRET-based functional imaging application. Moreover, when coupled with two-photon excitation, as proposed here, our technique should be well adapted to thick tissue imaging, thereby benefiting the in-vivo imaging community. PUBLIC HEALTH RELEVANCE: Fluorescence resonance energy transfer (FRET) is a powerful research technique to image cellular function in tissue. We propose to improve FRET signal generation with the use of ultrasound. This will benefit any FRET-based imaging applications, and will be specifically adapted to in-vivo microscopy in thick tissue.

描述（由申请人提供）：荧光共振能量转移（FRET）可以用作纳米级分子标尺。当在成像应用中使用时，它是供体 - 受体分子构型的高度敏感记者。在大多数情况下，用标准的一光激发使用FRET。通常会因供体和受体流血的问题而阻碍，将FRET扩展到两光子激发，这迫使技术复杂的光谱构成算法或荧光寿命测量值的要求。我们建议通过两光启动激发对FRET的检测进行大量简化。 我们的目标是建立使用超声波调节FRET信号的可行性。由于FRET对纳米级的供体对供体对之间的距离很敏感，因此声波产生的机械压缩和张力可能会调节FRET信号，作为将FRET与供体和受体出血的有用的签名，这是一种多光量特殊问题。 特别是，我们建议进行初步实验，以确定超声调节品格的前提。我们将研究两种类型的样品：组织中（或类似组织样环境）中的遗传编码的FRET探针，并在溶液中标记了微泡。在前一种情况下，我们将调查我们的技术在C.Elegans中的体内成像中的应用。 FRET是功能成像的强大工具。我们认为，混合超声波技术将显着改善FRET信号的隔离，因此对任何基于FRET的功能成像应用有益。此外，当此处提出的那样，当与两光蛋白激发相结合时，我们的技术应很好地适应厚的组织成像，从而使体内成像社区受益。 公共卫生相关性：荧光共振能量转移（FRET）是一种强大的研究技术，可在组织中图像细胞功能。我们建议通过使用超声来改善FRET信号的产生。这将受益于任何基于FRET的成像应用，并将专门适用于厚组织中的体内显微镜。