CAREER: Nanoscale sensing and imaging using computational single-molecule nanoscopy

职业：利用化学探针、计算和显微镜观察细胞膜内单个分子之间的相互作用细胞膜将其内部内容物（如细胞器、DNA和蛋白质）与环境分离，从而控制化学信号（如分子）如何发送到其他细胞并从其他细胞接收。由于这些膜的尺寸小、波动快，现有的成像技术很难将其纳米尺度的“皱纹”和粗糙度可视化。该项目将开发一种称为计算单分子纳米镜的协同方法，将光学硬件、图像处理软件和分子传感器结合起来，以纳米级分辨率对活细胞进行成像。提出的技术将可视化膜内的电和化学环境，控制它们如何工作。最终，这种方法将使科学家能够研究膜内的纳米级结构如何影响分子在膜上的运输，这可能对设计靶向和杀死癌细胞的纳米药物有用。PI将与圣路易斯科学中心和华盛顿大学SPECTRA学生小组合作，促进公众对这项研究的科学和技术理解。参与研究计划的本科生和研究生将获得应用物理学、光学、光谱学、估计理论、图像处理和生物学方面的广泛知识和各种技术技能。这些学生将为技术创新和科学发现方面的现代职业生涯做好独特而全面的准备。拟议的研究将开发一种集成的化学、光学和计算技术，称为计算单分子纳米镜，用于以纳米级分辨率感知和成像细胞膜的电学和化学特性。研究方法是：1)研究计算光学纳米显微镜测量纳米尺度信息的分辨率和灵敏度极限；2)量化荧光分子的性能，以感知其纳米环境的电学和化学性质；3)可视化细胞膜内的脂筏，以确定它们如何调节生物分子在细胞膜上的运输。这项工作的优点在于创新地利用荧光分子与光学显微镜之间的协同作用。与目前的方法简单地使用荧光分子作为报告生物分子位置的信标不同，快速扩散分子发出的荧光将被用来测量纳米环境的极性和流动性。为了利用这些分子报告的信息，新的光学系统和图像处理算法将共同设计，以最大限度地提高存在噪声的纳米尺度测量的精度和灵敏度。感知细胞膜的纳米级特性将为细胞间通讯提供新的见解：电、化学和机械信号如何在细胞之间和细胞膜之间传播。

CAREER: Visualizing the Interactions between Individual Molecules within Cell Membranes Using Chemical Probes, Computation, and MicroscopyThe membrane of a cell separates its internal contents, such as organelles, DNA, and proteins, from its environment and therefore controls how chemical signals (e.g., molecules) are sent to and received from other cells. Existing imaging technologies struggle to visualize the nanoscale "wrinkles" and roughness of these membranes due to their small sizes and fast fluctuations. This project will develop a synergistic approach called computational single-molecule nanoscopy that combines optical hardware, image processing software, and molecular sensors for imaging living cells with nanoscale resolution. The proposed technology will visualize the electrical and chemical environments within membranes that govern how they work. Ultimately, this approach will enable scientists to study how nanoscale structures within the membrane influence how molecules are transported across the membrane, which could be useful for the design of nanomedicines that target and kill cancer cells. The PI will collaborate with the Saint Louis Science Center and the Washington University SPECTRA student group to promote the public's scientific and technological understanding of this research. Undergraduate and graduate students involved in the research program will obtain broad knowledge and diverse technical skills in applied physics, optics, spectroscopy, estimation theory, image processing, and biology. These students will receive a unique and comprehensive preparation for modern careers in technology innovation and scientific discovery.The proposed research will develop an integrated chemical, optical, and computational technology, termed computational single-molecule nanoscopy, for sensing and imaging the electrical and chemical properties of cell membranes with nanoscale resolution. The research approach is 1) to investigate the resolution and sensitivity limits of computational optical nanoscopy for measuring nanoscale information; 2) to quantify the performance of fluorescent molecules for sensing the electrical and chemical properties of their nano-environments; and 3) to visualize lipid rafts within cell membranes to determine how they regulate the trafficking of biomolecules across the membrane. The merit of the proposed work lies in innovatively exploiting the synergy between fluorescent molecules and optical microscopes. Rather than simply using fluorescent molecules as beacons that report a biomolecule's location, as current methods do now, the fluorescence emitted by rapidly-diffusing molecules will be used to measure the polarity and fluidity of a nano-environment. To harness the information reported by these molecules, new optical systems and image processing algorithms will be jointly designed to maximize the precision and sensitivity of nanoscale measurements in the presence of noise. Sensing the nanoscale properties of cell membranes will provide new insight into intercellular communication: how electrical, chemical, and mechanical signals propagate between cells and across their membranes.