<> "The repository administrator has not yet configured an RDF license."^^ . <> . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy"^^ . "Small-molecule fluorophores have been recognized as efficient fluorescent reporters\r\nin the visualisation of living cells by super-resolution fluorescence microscopy. Much\r\nof the ongoing research focuses on positively charged fluorophores derived from rhodamine\r\ncores, which show good photostability, live cell permeability and brightness,\r\nbut suffer from undesired off-targeting and accumulation in certain cell compartments,\r\nmost typically in mitochondria. Furthermore, their derived photoactivatable labels often\r\nrely photocleavable caging groups as bulky nitrobenzene-type groups or α-diazoketones,\r\nwhich may generate toxic or non-fluorescent by-products.\r\nNegatively charged fluorophores derived from fluorescein present a valuable alternative\r\nwith no off-targeting and accumulation. Fluorescein, one of the oldest synthetic\r\nfluorophores, is a brightly emitting fluorophore, but suffer from low photostability and\r\ncell membrane-impermeability compared to rhodamines. Therefore, the rational design\r\nof negatively charged small-molecule fluorophores with improved photostability and cell\r\nmembrane-permeability is desired. Furthermore, photoactivatable fluorescein analogs\r\nmay rely on photocleavable groups less bulky than the nitrobenzene-types, which favor\r\nthe generation of non-toxic by-products, and hence useful for super-resolution microscopy.\r\nWithin this thesis, the chemical design and synthetic methods to access\r\nthese fluorophores are discussed and the photophysical properties and application in\r\nlive-cell super- resolution fluorescence microscopy examined.\r\nAs a result of this work, N-cyanorhodamines have been identified as bathochromically\r\nshifted fluorescein analogs with improved photostability and cell-permeability.\r\nThree different synthetic methods have been developed to access these fluorophores\r\nfrom known aryl triflates or iodides. Unlike tetramethylrhodamine, N-cyanorhodamines\r\ndemonstrate no off-targeting affinity. Based on these fluorophores, fluorescent ligands\r\nfor self-labeling protein tags HaloTag and SNAP-tag have been prepared and applied\r\nin combination with commercially available labels in multi-color confocal and stimulated\r\nemission depletion microscopy.\r\nThe second part of this thesis explores several different strategies to access photoactivatable\r\nfluorescein analogs bearing no or smallest possible photocleavable group. Protection\r\nof negatively charged fluorophores with photocleavable groups resulted in Ntetrazinylrhodamines\r\nand mercaptofluorans as unstable or non-photoactivatable compounds.\r\nOn the other hand, a synthetic route towards 3,6-dihydroxyxanthones on previously\r\ndescribed caging-group-free photoactivatable PaX dyes has been established.\r\nEfficient photoactivation of these compounds indicated their potential for future application\r\nin single-molecule localisation imaging and as enzymatic sensor activity probes.\r\nIn conclusion, it was demonstrated that negatively charged fluorescein analogs represents\r\na valuable addition to the toolkit of live cell-compatible fluorescent dyes. Within\r\nthis work, N-cyanorhodamines as highly photostable and cell membrane-permeable\r\nfluorescein derivatives and hydroxy analogs of PaX dyes as photoactivatable analogs\r\nof fluoresceins have been prepared. The application of such dyes as fluorophores in\r\nlive cell imaging has been demonstrated."^^ . "2024" . . . . . . . "Lukas"^^ . "Heynck"^^ . "Lukas Heynck"^^ . . . . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (PDF)"^^ . . . "DissertationHeynckLukas.pdf"^^ . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (Other)"^^ . . . . . . "indexcodes.txt"^^ . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (Other)"^^ . . . . . . "lightbox.jpg"^^ . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (Other)"^^ . . . . . . "preview.jpg"^^ . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (Other)"^^ . . . . . . "medium.jpg"^^ . . . "Design and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy (Other)"^^ . . . . . . "small.jpg"^^ . . "HTML Summary of #32463 \n\nDesign and synthesis of anionic and photoactivatable fluorophores for super-resolution microscopy\n\n" . "text/html" . .