Optical microscopes are routinely employed for imaging live cell dynamics. Until recently, conventional optical microscopes lacked the ability to resolve spatial features significantly smaller than the wavelength of light. This kept the structure and dynamics of a vast array of biological processes hidden. Understanding the spatial organization and temporal dynamics of nanoscale molecular assemblies is critical to developing a comprehensive understanding of biology. In recent years, super-resolution (SR) microscopes have enabled routine live cell imaging at spatial resolutions <50nm. These new tools produced discoveries that challenged multiple paradigms of intracellular processes. Because optical scattering severely distorts SR methods, the SR imaging revolution has failed to be translated deep into scattering tissue. Yet it is well known that the behavior of cells in tissues and tumors deviates strongly from the behavior of 2D cell cultures. Here we present a new approach to optical SR imaging with spatial frequency modulated imaging that is, in principle, capable of providing unrestricted spatial resolution deep in live animal tissues. A broad illumination bandwidth homogenizes speckle that would otherwise be accumulated by the spatiotemporally structured illumination light, thereby preventing the speckle from distorting the image formation process. Further, scattering of the fluorescent light emitted from the object does not impact the quality of the measured image. We detail the principles of this SR imaging method and present both analytical and numerical calculations that test these concepts. Such discoveries will likely drive an improvement in our understanding of biology and disease.
Jeffrey J. Field, Patrick Stockton, and Randy A. Bartels, "Unrestricted superresolution imaging in scattering tissues with spatial frequency modulated imaging
(Conference Presentation)," Proc. SPIE 10500, Single Molecule Spectroscopy and Superresolution Imaging XI, 105000I (Presented at SPIE BiOS: January 28, 2018; Published: 14 March 2018); https://doi.org/10.1117/12.2290271.5751635031001.
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