Polar Red-Emitting Rhodamine Dyes with Reactive Groups: Synthesis, Photophysical Properties, and Two-Color STED Nanoscopy Applications
2014 | journal article; research paper. A publication with affiliation to the University of Göttingen.
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Polar Red-Emitting Rhodamine Dyes with Reactive Groups: Synthesis, Photophysical Properties, and Two-Color STED Nanoscopy Applications
Kolmakov, K. ; Wurm, C. A. ; Meineke, D. N. H.; Göttfert, F. ; Boyarskiy, V. P. ; Belov, V. N. & Hell, S. (2014)
Chemistry - A European Journal, 20(1) pp. 146-157. DOI: https://doi.org/10.1002/chem.201303433
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- Authors
- Kolmakov, Kirill ; Wurm, Christian Andreas ; Meineke, Dirk N. H.; Göttfert, Fabian ; Boyarskiy, Vadim P. ; Belov, Vladimir N. ; Hell, Stefan
- Abstract
- The synthesis, reactivity, and photophysical properties of new rhodamines with intense red fluorescence, two polar residues (hydroxyls, primary phosphates, or sulfonic acid groups), and improved hydrolytic stability of the amino-reactive sites (NHS esters or mixed N-succinimidyl carbonates) are reported. All fluorophores contain an N-alkyl-1,2-dihydro-2,2,4-trimethylquinoline fragment, and most of them bear a fully substituted tetrafluoro phenyl ring with a secondary carboxamide group. The absorption and emission maxima in water are in the range of 635-639 and 655-659nm, respectively. A vastly simplified approach to red-emitting rhodamines with two phosphate groups that are compatible with diverse functional linkers was developed. As an example, a phosphorylated dye with an azide residue was prepared and was used in a click reaction with a strained alkyne bearing an N-hydroxysuccinimid (NHS) ester group. This method bypasses the undesired activation of phosphate groups, and gives an amphiphilic amino-reactive dye, the solubility and distribution of which between aqueous and organic phases can be controlled by varying the pH. The presence of two hydroxyl groups and a phenyl ring with two carboxyl residues in the dyes with another substitution pattern is sufficient for providing the hydrophilic properties. Selective formation of a mono-N-hydroxysuccinimidyl ester from 5-carboxy isomer of this rhodamine is reported. The fluorescence quantum yields varied from 58 to 92% for free fluorophores, and amounted to 18-64% for antibody conjugates in aqueous buffers. The brightness and photostability of these fluorophores facilitated two-color stimulated emission depletion (STED) fluorescence nanoscopy of biological samples with high contrast and minimal background. Selecting a pair of fluorophores with absorption/emission bands at 579/609 and 635/655nm enabled two-color channels with low cross-talk and negligible background at approximately 40nm resolution.
- Issue Date
- 2014
- Journal
- Chemistry - A European Journal
- ISSN
- 0947-6539
- eISSN
- 1521-3765
- Language
- English