title: RESOLFT nanoscopy with photoswitchable organic fluorophores.
abstract: Far-field optical nanoscopy has been widely used to image small objects with sub-diffraction-limit spatial resolution. Particularly, reversible saturable optical fluorescence transition (RESOLFT) nanoscopy with photoswitchable fluorescent proteins is a powerful method for super-resolution imaging of living cells with low light intensity. Here we demonstrate for the first time the implementation of RESOLFT nanoscopy for a biological system using organic fluorophores, which are smaller in size and easier to be chemically modified. With a covalently-linked dye pair of Cy3 and Alexa647 to label subcellular structures in fixed cells and by optimizing the imaging buffer and optical parameters, our RESOLFT nanoscopy achieved a spatial resolution of ~74 nm in the focal plane. This method provides a powerful alternative for low light intensity RESOLFT nanoscopy, which enables biological imaging with small organic probes at nanoscale resolution.
authors.affiliation: Department of Biophysics and Chemical Biology, Seoul National University, Seoul 08826, Korea.
authors.forename: Jiwoong
authors.initials: J
authors.lastname: Kwon
authors.affiliation: Department of Biophysics and Chemical Biology, Seoul National University, Seoul 08826, Korea.
authors.forename: Jihee
authors.initials: J
authors.lastname: Hwang
authors.affiliation: Department of Chemistry, Seoul National University, Seoul 08826, Korea.
authors.forename: Jaewan
authors.initials: J
authors.lastname: Park
authors.affiliation: Department of Chemistry, Seoul National University, Seoul 08826, Korea.
authors.forename: Gi Rim
authors.initials: GR
authors.lastname: Han
authors.affiliation: Howard Hughes Medical Institute, Urbana, Illinois 61801, USA.|Department of Physics and Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
authors.forename: Kyu Young
authors.initials: KY
authors.lastname: Han
authors.affiliation: Department of Biophysics and Chemical Biology, Seoul National University, Seoul 08826, Korea.|Department of Chemistry, Seoul National University, Seoul 08826, Korea.
authors.forename: Seong Keun
authors.initials: SK
authors.lastname: Kim
mesh_terms: D000818:Animals; D002021:Buffers; D002232:Carbocyanines / Q000737:chemistry; D019281:Dimerization; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D021621:Imaging, Three-Dimensional; D008027:Light; D008297:Male; D051379:Mice; D015335:Molecular Probes / Q000737:chemistry; D041681:NIH 3T3 Cells; D036103:Nanotechnology / Q000379:methods*; D009930:Organic Chemicals / Q000737:chemistry*; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: C569686:Alexa Fluor 647; D002021:Buffers; D002232:Carbocyanines; D005456:Fluorescent Dyes; D015335:Molecular Probes; D009930:Organic Chemicals; C089187:cyanine dye 3