@phdthesis{Waeldchen2020,
  author    = {W{\"a}ldchen, Felix},
  title     = {3D Single Molecule Imaging In Whole Cells Enabled By Lattice Light-Sheet Illumination},
  doi       = {10.25972/OPUS-20711},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-207111},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2020},
  abstract  = {Single molecule localization microscopy has seen a remarkable growth since its first experimental implementations about a decade ago. Despite its technical challenges, it is already widely used in medicine and biology and is valued as a unique tool to gain molecular information with high specificity. However, common illumination techniques do not allow the use of single molecule sensitive super-resolution microscopy techniques such as direct stochastic optical reconstruction microscopy (dSTORM) for whole cell imaging. In addition, they can potentially alter the quantitative information. In this thesis, I combine dSTORM imaging in three dimensions with lattice lightsheet illumination to gain quantitative molecular information from cells unperturbed by the illumination and cover slip effects. Lattice light-sheet illumination uses optical lattices for beam shaping to restrict the illumination to the detectable volume. I describe the theoretical background needed for both techniques and detail the experimental realization of the system as well as the software that I developed to efficiently evaluate the data. Eventually, I will present key datasets that demonstrate the capabilities of the developed microscope system with and without dSTORM. My main goal here was to use these techniques for imaging the neural cell adhesion molecule (NCAM, also known as CD56) in whole cells. NCAM is a plasma membrane receptor known to play a key role in biological processes such as memory and learning. Combining dSTORM and lattice light-sheet illumination enables the collection of quantitative data of the distribution of molecules across the whole plasma membrane, and shows an accumulation of NCAM at cell-cell interfaces. The low phototoxicity of lattice light-sheet illumination further allows for tracking individual NCAM dimers in living cells, showing a significant dependence of its mobility on the actin skeleton of the cell.},
  subject      = {Einzelmolek{\"u}lmikroskopie},
  language  = {en}
}