Three-Dimensional (3D) Acoustofluidic Scanning Nanoscope with Super Resolution and Large Field of View

项目总结 在过去的二十年里，已经开发了许多“超分辨率”3D成像技术， 使研究人员能够观察到以前传统的、 衍射限制成像技术。在纳米尺度上可视化细胞和亚细胞结构的能力 揭示了对各种生物过程的关键见解。尽管已经取得了令人印象深刻的进展 在3D超分辨率成像技术的发展中，研究人员经常被迫接受权衡 在分辨率、视野、速度和3D成像技术的易用性方面。最近，我们有 开发了一种声流扫描纳米显微镜，它可以同时实现超分辨率和 大视场2D成像。在这个R01项目中，我们将开发和验证3D声流扫描 具有以下特点的纳米显微镜：(1)横向和轴向分辨率为~50的超分辨率成像 纳米和~120纳米：提出的3D成像方法将达到四倍的分辨率 比共焦显微镜更好，使光学成像更详细的内部结构 多种亚细胞结构；(2)大视场(约1,100×1,100微米)：传统光学 成像方法以降低分辨率为代价实现高通量成像，反之亦然。通过利用 声学同时操纵多个微球透镜，提出的成像方法将解决 这一长期存在的技术障碍是在大视场成像的同时保持优越的横向和轴向成像 分辨率；(3)成像速度比共焦显微镜快10倍：快速z堆叠 通过使用表面声波来实现比共焦显微镜快10倍的速度 以精确、可控的方式扫描样品体积上的微球阵列；(4)无缝 连接到传统光学显微镜以便于使用：我们的设备可以无缝连接 在不修改光学设置的情况下，这可以显著地减少 成本和操作的复杂性。具有上述优点，提出的3D声流控 扫描纳米技术有可能大大超过该领域的当前标准，并 解决许多未得到满足的需求。我们将通过成像3D纳米棒样品和细胞器来验证其性能 活的HeLa细胞。在这方面，我们的目标是展示我们的3D声流控技术的深远潜力 扫描纳米技术，使从亚细胞成像到 3D神经活动的可视化。

PROJECT SUMMARY Over the past two decades, a number of “super-resolution” 3D imaging technologies have been developed, enabling researchers to observe nanoscale biological structures that were previously invisible to traditional, diffraction-limited imaging techniques. The ability to visualize cellular and subcellular structures at the nanoscale has revealed key insights into a variety of biological processes. Although impressive progress has been made in the development of 3D super-resolution imaging techniques, researchers are often forced to accept a tradeoff in terms of the resolution, field-of-view, speed, and ease of use of their 3D imaging technique. Recently, we have developed an acoustofluidic scanning nanoscope that can simultaneously achieve both super-resolution and large field-of-view imaging in 2D. In this R01 project, we will develop and validate a 3D acoustofluidic scanning nanoscope with the following features: (1) Super-resolution imaging with lateral and axial resolutions of ~50 nm and ~120 nm, respectively: The proposed 3D imaging method will achieve a resolution that is four times better than that from a confocal microscope, which makes the optical imaging of more detailed inner architecture of many subcellular structures possible; (2) Large field-of-view (~1,100×1,100 µm2): Conventional optical imaging methods achieve high-throughput imaging at the cost of reduced resolution and vice versa. By utilizing acoustics to simultaneously manipulate multiple microsphere lenses, the proposed imaging method will solve this long-standing technical barrier for large field-of-view imaging while maintaining superior lateral and axial resolution; (3) Imaging speed 10 times faster than that from a confocal microscope: Rapid z-stacking at a speed 10 times faster than that of a confocal microscope can be achieved by using surface acoustic waves to scan an array of microspheres across the sample volume in a precise, controllable manner; (4) Seamless connection to a conventional optical microscope for ease of use: Our device can be seamlessly connected to a conventional optical microscope without modification of the optical setup, which can significantly reduce the cost and the complexity of operation. With the aforementioned advantages, the proposed 3D acoustofluidic scanning nanoscope technology has the potential to significantly exceed current standards in the field and address many unmet needs. We will validate its performance by imaging 3D nanorod samples and the organelles of live HeLa cells. In this regard, we aim to demonstrate the far-reaching potential of our 3D acoustofluidic scanning nanoscope technology to enable improved research in areas ranging from subcellular imaging to the visualization of 3D neural activity.