Brillouin Microscope for Biomedical Research

这一提议的动机是需要评估微观的粘弹性属性，这些属性已经出现在 一种强大的生物标志物，可用于多种疾病，如癌症、动脉粥样硬化、镰状细胞病等，但 也已被确定为许多生物过程的驱动力，例如致癌， 血管生成、形态发生等。再生医学新生物材料的出现也呼吁 以更好地了解生物力学细胞水平的相互作用。 在过去，对组织弹性特性的评估大多局限于使用 超声和磁共振成像以及使用以下任一光学技术的纳米接触评估 镊子或原子力显微镜仪器，这为我们更好地理解 细胞和组织的粘弹性特性及其对生物医学研究的重要性。同时，它也是 现在意识到在能够非侵入性评估的仪器方面有很大的技术差距 具有足够高空间分辨率、高灵敏度和高分辨率的微观尺度的粘弹性特性 速度。最近，光学相干弹性成像被成功地开发来评估弹性性能 15-100𝜇𝑚范围内的组织。理想情况下，这样的工具应该与现有的 使用荧光和拉曼显微镜系统的仪器，以提供额外的能力 那些。布里渊显微镜正在成为非侵入性生物医学成像的有力工具。开发它 成为生物医学研究的强大工具，并有可能应用于临床，被认为是 这项提议的首要目标。 通过这项赠款申请，将采取两项战略。第一种方法是依靠自发的 布里渊显微镜，在使用上更简单，并且经过相对较小的修改，可以实现为 用于大型生物医学社区的现有商用荧光或拉曼显微镜的一种选择。 第二种策略是利用非线性布里渊光谱和显微技术来提高 信号和数据采集速度高达惊人的5个数量级。这种方法利用超短波 脉冲激发，与多光子荧光显微镜、二次和三次谐波完全兼容 显微镜和相干反斯托克斯拉曼显微镜。非线性布里渊的另一个好处 显微镜是改进的切片能力。总体战略是设计、建造和表征 这两个显微镜是平行的，因为它们中的每一个都为特定的一组应用提供了不同的优点， 并展示它们为生物相关系统成像的能力，以成像细胞生长和 斑马鱼胚胎发育对局部粘弹性环境的响应和图像发育 受精后72小时。

This proposal is motivated by the need to assess microscopic viscoelastic properties, which have emerged as a powerful biomarker for a number of diseases, such as cancer, atherosclerosis, sickle cell disease, etc., but also have been identified as a driving force for many biological processes, such as carcinogenesis, angiogenesis, morphogenesis, etc. The emergence of novel biomaterials for regenerative medicine also calls for a better understanding of biomechanical cellular-level interactions. In the past, assessment of elastic properties of tissues was mostly limited to large-scale imaging using ultrasound and magnetic resonant imaging and to nanoscopic contact assessment using either optical tweezers or atomic force microscopy instruments, which paved the way to our better understanding of viscoelastic properties of cells and tissues and their importance for biomedical research. In the same time, it is now realized that there is a substantial technology gap in instrumentation capable of assessing non-invasively viscoelastic properties on a microscopic scale with high enough spatial resolution, high sensitivity and high speed. Recently, optical coherence elastography was successfully developed to assess elastic properties of tissues on the scale of 15-100 𝜇𝑚. Ideally, such an instrument should be fully compatible with existing instrumentation using fluorescence and Raman microscopy systems to provide an additional capability to those. Brillouin microscopy is emerging as a powerful tool for non-invasive biomedical imaging. Developing it into a powerful instrument for biomedical research and, potentially, clinical applications is considered to be the overarching goal of this proposal. Two strategies will be pursued through this grant application. The first approach is relying on spontaneous Brillouin microscopy, which is simpler in use, and, with relatively minor modifications, can be implemented as an option in already existing commercial fluorescent or Raman microscopes for a large biomedical community. The second strategy is to utilize nonlinear Brillouin spectroscopy and microscopy to boost the efficiency of the signal and data acquisition rate by astonishing 5 orders of magnitude. This methodology utilizes ultrashort pulse excitation and is fully compatible with multiphoton fluorescence microscopy, second- and third-harmonic microscopies and coherent anti-Stokes Raman microscopy. An additional benefit of nonlinear Brillouin microscopy is improved sectioning capabilities. The overall strategy is to design, construct and characterize both microscopes in parallel, since each of those offers distinct advantages for a particular set of applications, and to demonstrate their imaging capabilities for biologically relevant systems to image cells growth and development in response to a local viscoelastic environment and image developing zebrafish embryo during the first 72 hours post fertilization.