Far‐Red Emitting Fluorescent Dyes for Optical Nanoscopy: Fluorinated Silicon–Rhodamines (SiRF Dyes) and Phosphorylated Oxazines |
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Authors: | Dr Kirill Kolmakov Elke Hebisch Dr Thomas Wolfram Lars A Nordwig Dr Christian A Wurm Dr Haisen Ta Dr Volker Westphal Dr Vladimir N Belov Prof Stefan W Hell |
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Institution: | Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 G?ttingen (Germany) |
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Abstract: | Far‐red emitting fluorescent dyes for optical microscopy, stimulated emission depletion (STED), and ground‐state depletion (GSDIM) super‐resolution microscopy are presented. Fluorinated silicon–rhodamines (SiRF dyes) and phosphorylated oxazines have absorption and emission maxima at about λ≈660 and 680 nm, respectively, possess high photostability, and large fluorescence quantum yields in water. A high‐yielding synthetic path to introduce three aromatic fluorine atoms and unconventional conjugation/solubilization spacers into the scaffold of a silicon–rhodamine is described. The bathochromic shift in SiRF dyes is achieved without additional fused rings or double bonds. As a result, the molecular size and molecular mass stay quite small (<600 Da). The use of the λ=800 nm STED beam instead of the commonly used one at λ=750–775 nm provides excellent imaging performance and suppresses re‐excitation of SiRF and the oxazine dyes. The photophysical properties and immunofluorescence imaging performance of these new far‐red emitting dyes (photobleaching, optical resolution, and switch‐off behavior) are discussed in detail and compared with those of some well‐established fluorophores with similar spectral properties. |
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Keywords: | dyes/pigments fluorescence structure– activity relationships super‐resolution microscopy synthesis design |
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