RESUMO
Within the last decade, super-resolution methods that surpass the diffraction limit of light microscopy have provided invaluable insight into a variety of biological questions. Each of these approaches has inherent advantages and limitations, such that their combination is a powerful means to transform them into versatile tools for the life sciences. Here, we report the development of a combined SIM and STORM setup that maintains the optimal resolution of both methods and which is coupled to image registration to localize biological structures in 3D using multicolor labeling. We utilized this workflow to determine the localization of Bld12p/CrSAS-6 in purified basal bodies of Chlamydomonas reinhardtii with utmost precision, demonstrating its usefulness for accurate molecular mapping in 3D.
RESUMO
We introduce far-red, fluorogenic probes that combine minimal cytotoxicity with excellent brightness and photostability for fluorescence imaging of actin and tubulin in living cells. Applied in stimulated emission depletion (STED) microscopy, they reveal the ninefold symmetry of the centrosome and the spatial organization of actin in the axon of cultured rat neurons with a resolution unprecedented for imaging cytoskeletal structures in living cells.
