CAREER: Three-dimensional super-resolution light microscopy of thick, unprocessed biological samples

孟菲斯大学被授予为未染色的细胞、组织和器官开发一种创新的成像系统。目前的成像系统不能对厚样品进行高分辨率成像。该项目将通过开发一种仪器来克服这一限制，该仪器将提供高分辨率的厚样品成像。该项目将结合研究和教育，以激发对光学工程的兴趣，为学生提供一套独特的技能，以在他们的专业生涯中取得成功。光学工程是专注于光学和光子学的发展和应用的研究领域，包括仪器仪表。在这个项目中，五项教育活动的重点是传授光学工程知识。这些教育活动包括：(1)在线光学工程证书；(2)高级光学显微技术暑期工作坊；(3)在洛桑学院创建STEM俱乐部，让高中生可以开发在科学课堂上通常不可能完成的科学项目；(4)与粉色故宫博物院合作，让公众接触光学和显微镜；以及(5)为本科生提供暑期实习，让他们接触研究活动和新的职业道路。这个项目有几个更广泛的影响。最重要的更广泛的影响是增强了生物学研究的成像基础设施，并扩大了我们对细胞行为的知识。另一个重要的更广泛的影响是本科生和研究生的就业机会增加。在这个项目中，PI将使用YouTube平台创建研究、教育和科学外展视频。这些视频将提供构建成像系统以及使用光学工具对其进行对准的实践演示。参与该项目的学生将参与这些视频，使他们对主题有更深入的理解，并提高他们的沟通技能。这些视频将以英语和西班牙语音频提供，以支持美国各地拉美裔学生的教育成长。现有的光学显微镜方法无法实现对厚(50μm)生物样本的三维(3D)超分辨率(SR)成像而不染色。这一局限性导致我们对未染色标本的3D细胞形状和行为发生的动态变化的理解有很大差距。这个项目将使生物研究人员能够提高他们对与细胞生长和动力学有关的大量生物学问题的知识，以及研究厚组织切片中的细胞行为。拟议的成像系统的智能优点包括在硬件和计算方法方面的新进展。该系统的评估将使用校准的制造对象和相关的生物标本，包括人神经母细胞瘤细胞、原代小鼠干细胞和小鼠胚泡。参与这一项目的跨学科团队将系统地将使用拟议系统获得的图像与现有方法获得的图像进行比较，包括共聚焦显微镜、狮心FX系统和NanoLive平台。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

An award is made to the University of Memphis to develop an innovative imaging system for unstained cells, tissues, and organs. Current imaging systems cannot image thick samples with high resolution. This project will overcome this limitation by developing an instrument that will provide thick-sample imaging with high resolution. This project will integrate research and education to stimulate interest in Optical Engineering, providing students with a unique set of skills for succeeding in their professional careers. Optical Engineering is the field of study that focuses on the development and application of optics and photonics, including instrumentation. In this project, five educational activities are focused on transferring the knowledge of Optical Engineering. These educational activities include the following: (1) an online Optical Engineering Certificate; (2) a Summer Workshop in Advanced Optical Microscopic Techniques; (3) the creation of a STEM club in Lausanne Collegiate School so high-school students can develop science projects that are generally not possible in science classrooms; (4) a partnership with the Pink Palace Museum to expose the public to optics and microscopy; and (5) summer internships to undergraduate students, exposing them to research activities and new career paths. This project has several broader impacts. The most important broader impact is the enhancement of the imaging infrastructure for biological research and expanding our knowledge of cell behavior. Another significant broader impact is the increase in career opportunities for undergraduate and graduate students. During this project, the PI will create research, education, and scientific outreach videos using the YouTube platform. These videos will provide hands-on demonstrations of building imaging systems and the use of optical tools to align them. Students involved in this project will participate in these videos, allowing them to develop a deeper understanding of the topic and improving their communication skills. The videos will be available in English and Spanish audio to support the educational growth of Hispanic students across the United States.Available methods of light microscopy do not enable three-dimensional (3D) super-resolution (SR) imaging of thick (50 μm) biological samples without staining. This limitation results in a significant gap in our understanding of dynamic changes occurring in the 3D cell-shape and behavior of unstained specimens. This project will enable biological investigators to advance their knowledge of a large number of biological questions relating to cell growth and dynamics as well as for investigating cell behavior in thick tissue slices. The intellectual merit of the proposed imaging system includes novel advances in hardware and computational methods. The evaluation of the proposed system will be performed using calibrated manufactured objects and relevant biological specimens including human neuroblastoma cells, primary murine stem cells, and murine blastocysts. The interdisciplinary team involved in this project will systematically compare images obtained with the proposed system with images from existing methods including confocal microscopy, the Lionheart FX system, and the NanoLive platform.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.