Collaborative Research: QSTORM: Switchable Quantum Dots and Adaptive Optics for Super-Resolution Imaging

[52623]冬，张晓东，张晓东。（牵头项目）合作研究：QSTORM：可切换量子点和超分辨率成像的自适应光学。然而，在低于光学显微镜衍射极限（~200 nm）的分辨率下观察活细胞中的生物结构和过程仍然极具挑战性。近年来，为了提高光学荧光的分辨率，引入了几种超分辨率技术，据报道，静态和动态分辨率分别达到~20 nm和~60 nm。然而，由于荧光探针的局限性以及组织中的光学像差和光散射造成的困难，这些技术尚未转化为活细胞。因此，研究人员必须从固定标本的图像中推断出生活状态的信息。该项目提出了一种新的超分辨率成像技术：QSTORM，它将用户控制的可切换量子点（QDs）与专门的基于计算机的算法（STORM）和自适应光学相结合，以增强图像。QSTORM将首次使活细胞成像的分辨率优于或可与其他超分辨率技术相媲美。QSTORM将在两个模型系统中进行评估：斑马鱼肌肉肌丝的结构和功能以及果蝇神经元囊泡的细胞内运输。正常的肌肉功能依赖于肌肉组织的高度组织化的多尺度结构。QSTORM可以同时对同一样品中的肌丝、肌节和整个肌肉细胞进行成像。同样，通过囊泡进行的轴突货物运输对神经元细胞的存活和功能至关重要。QSTORM将允许观察单个囊泡的运动和绘制使这种运输成为可能的潜在细胞骨架结构。此外，QSTORM团队将与波士顿的科学博物馆合作，通过科学教育计划、博物馆演示和基于网络的多媒体项目广泛地分享这项研究的结果。知识价值。如果完全成功，QSTORM将利用量子点和自适应光学的优越成像能力，在小于50纳米的超分辨率下进行活细胞成像。QSTORM将改变生物过程的成像，特别是那些涉及细胞骨架和运动蛋白的成像。在待研究的模型中，QSTORM将允许完整活肌肉的三维高分辨率成像，而无需透射电子显微镜（TEM）所需的破坏性处理，从而可能导致肌肉蛋白（如肌动蛋白、肌球蛋白和相关蛋白）如何相互作用的新假设。同样，QSTORM将首次允许在单纳米分辨率下沿整个轴突长度对神经元囊泡在完整运输周期中的运动进行成像，从而有可能改变目前对运输及其调控的基本分子机制的理解。更广泛的影响。QSTORM将为科学界提供一个强大的新型显微镜工具。这项研究不仅会产生非凡的图像，提供对基本生物过程的视觉洞察，而且更广泛的传播结果和教育活动将广泛推进亚细胞生物学的研究和培训。研究人员、学生、教育工作者和公众将从QSTORM产生的潜在非凡的可视化中受益。这项研究将被纳入广泛学科的研究生和本科生课程。与科学博物馆的合作将提供广泛获取和在全国传播的教育材料。拟议中的QSTORM网站将展示这些非凡的图像，作为一个生动的多媒体故事的一部分，讲述高风险、跨学科的科学和技术合作，以追求一个巨大的挑战。

1052623Winter, Jessica O. (lead PI)Collaborative Research: QSTORM: Switchable Quantum Dots and Adaptive Optics for Super-Resolution ImagingImaging is one of the most important tools in biology. However, observing biological structures and processes in living cells at a resolution below the diffraction limit of light microscopy (~200 nm) remains extremely challenging. Recently, several super-resolution techniques have been introduced to improve the resolution of optical fluorescence, with reported static and dynamic resolutions reaching ~20 nm and ~60 nm, respectively. However, these techniques have yet to be translated to the live cell because of difficulties caused by limitations of fluorescent probes and optical aberrations and light scattering in tissues. Thus researchers must extrapolate information from images of fixed specimens to the living state. This project proposes a new super-resolution imaging technology: QSTORM, which combines user-controlled, switchable quantum dots (QDs) with specialized computer-based algorithms (STORM) and adaptive optics to enhance images. QSTORM will, for the first time, enable imaging in living cells with a resolution superior or comparable to other super-resolution techniques. QSTORM will be evaluated in two models systems: the structure and function of muscle myofilaments in zebrafish and the intracellular transport of vesicles in fruit fly neurons. Normal muscle function depends on the highly organized multi-scale architecture of muscle tissue. QSTORM will enable simultaneous imaging of functioning myofilaments, sarcomeres, and whole muscle cells within the same sample. Similarly, axonal transport of cargo by vesicles is critical to the survival and function of neuronal cells. QSTORM will permit observation of the movements of individual vesicles and the mapping of the underlying cytoskeletal structures that enable this transport. Additionally, the QSTORM team will collaborate with the Museum of Science in Boston to share the results of this research broadly through science education programs, museum demonstrations, and Web-based multimedia projects.Intellectual merit. If fully successful, QSTORM will harness the superior imaging capabilities of quantum dots and adaptive optics for live cell imaging at a super-resolution of less than 50 nm. QSTORM will transform imaging of biological processes, particularly those involving the cytoskeleton and motor proteins. In the models to be studied, QSTORM will permit three-dimensional high resolution imaging of intact live muscle without the destructive processing required for transmission electron microscopy (TEM), thus potentially leading to new hypotheses of how muscle proteins such as actin, myosin, and associated proteins interact. Similarly, QSTORM will permit, for the first time, imaging the movements of neuronal vesicles over complete transport cycles along the entire length of the axon at single nanometer resolution, thus potentially transforming current understanding of the fundamental molecularmechanisms of transport and its regulation.Broader impacts. QSTORM will contribute a powerful new microscopy tool to the scientific community. Not only will this research produce extraordinary images that offer visual insight into fundamental biological processes, but also the broader dissemination of results and educational activities will widely advance subcellular biological research and training. Researchers, students, educators, and public audiences will benefit from the potentially extraordinary visualizations produced by QSTORM. This research will be incorporated into graduate and undergraduate courses in a wide-range of disciplines. Collaboration with the Museum of Science will provide broadly accessible and nationally disseminated educational materials. The proposed QSTORM Web site will present these extraordinary images as part of a lively multimedia story of high-risk, interdisciplinary scientific and technical collaboration in pursuit of a grand challenge.