Nonlinear optical spectroscopy and super-resolution microscopy

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Studies of Water Surface Chemistry Using Nonlinear Optical Spectroscopy       This proposal applies nonlinear optical spectroscopy to study environmentally- and industrially-relevant surface chemistry of water. Nonlinear optical spectroscopy, such as sum-frequency generation, is one of the few techniques available to measure the vibrational spectra of a buried liquid/solid interface. The vibrational spectra provide valuable information on the molecular composition and organization of water interfaces. This proposal will study heterogeneous ice formation at water/solid interfaces, aiming to understand ice cloud formation in the atmosphere, and the molecular adsorption kinetics at water/mineral interfaces, aiming to reduce the consumption of resources and environmental impacts of the oil sands development in Canada. Super-Resolution Microscopy: From Single-Molecule Spectroscopy to Protein Assemblies in Whole Cells       In the past several years, scientists have developed several approaches to overcome the diffraction limit and revolutionized optical microscopy. Single-molecule localization microscopy (SMLM) has provided an unprecedented spatial resolution of 10 nm. SMLM techniques activate and locate only a sparse subset of fluorophores at any given time. Because the images of each fluorophore is well separated spatially, the location of each fluorophore can be determined with a very high accuracy. A super-resolution image can be reconstructed using the x-, y-, and z- coordinates of fluorophores obtained from a series of diffraction-limited images. These 3D point clouds cannot be analyzed using traditional image analysis tools. We will employ machine learning to mimic the modulation transfer function of human eyes to analyze the 3D point clouds produced by SMLM. Since in many aspects humans outperform any pattern-recognition machines, this approach will lead to an effective protein cluster analysis tool without any user input.       Additionally, we propose to implant a sample stabilization system into a structured illumination microscope (SIM) to remove artifacts due to sample drift. Compared to SMLM, SIM has several advantages for 3D imaging. It has a much higher speed, uses a relatively low illumination power, and is easily available for multicolor imaging. The proposed SIM will enable us to obtain reliable super-resolution images for large cells, such as cardiac myocytes, which has a mean length of 100 µm and a width of 18 µm. The microscope will allow us to image protein organizations in whole cells and make exciting discoveries.