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Kirill Kolmakov

Bio: Kirill Kolmakov is an academic researcher from Max Planck Society. The author has contributed to research in topics: Rhodamine & Rhodamines. The author has an hindex of 13, co-authored 19 publications receiving 834 citations.

Papers
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Journal ArticleDOI
TL;DR: Novel rhodamine dyes excitable with 630 nm laser light and emitting at around 660 nm have been developed and Syntheses of lipophilic and hydrophilic derivatives starting from the same chromophore-containing scaffold are described.
Abstract: Fluorescent markers emitting in the red are extremely valuable in biological microscopy since they minimize cellular autofluorescence and increase flexibility in multicolor experiments. Novel rhodamine dyes excitable with 630 nm laser light and emitting at around 660 nm have been developed. The new rhodamines are very photostable and have high fluorescence quantum yields of up to 80 %, long excited state lifetimes of 3.4 ns, and comparatively low intersystem-crossing rates. They perform very well both in conventional and in subdiffraction-resolution microscopy such as STED (stimulated emission depletion) and GSDIM (ground-state depletion with individual molecular return), as well as in single-molecule-based experiments such as fluorescence correlation spectroscopy (FCS). Syntheses of lipophilic and hydrophilic derivatives starting from the same chromophore-containing scaffold are described. Introduction of two sulfo groups provides high solubility in water and a considerable rise in fluorescence quantum yield. The attachment of amino or thiol reactive groups allows the dyes to be used as fluorescent markers in biology. Dyes deuterated at certain positions have narrow and symmetrical molecular mass distribution patterns, and are proposed as new tags in MS or LC-MS for identification and quantification of various substance classes (e.g., amines and thiols) in complex mixtures. High-resolution GSDIM images and live-cell STED-FCS experiments on labeled microtubules and lipids prove the versatility of the novel probes for modern fluorescence microscopy and nanoscopy.

228 citations

Journal ArticleDOI
28 Sep 2012
TL;DR: Four novel red fluororescent dyes are reported on, including the first phosphorylated dye, created by combining a rigidized rhodamine backbone with various polar groups, which exhibit large fluorescence quantum yields and improved NHS ester stability.
Abstract: In optical microscopy, most red-emitting dyes provide only moderate performance due to unspecific binding, poor labeling efficiency, and insufficient brightness. Here we report on four novel red fluororescent dyes, including the first phosphorylated dye, created by combining a rigidized rhodamine backbone with various polar groups. They exhibit large fluorescence quantum yields and improved NHS ester stability. While these fluorophores are highly suitable for fluorescence microscopy in general, they excel in stimulated emission depletion (STED) microscopy, providing < 25 nm spatial resolution in raw images of cells.

104 citations

Journal ArticleDOI
TL;DR: This work presents a general strategy to covalently link a synthetic organic fluorophore simultaneously to a photostabilizer and biomolecular target via unnatural amino acids, and is convinced that the presented scaffolding strategy and the improved characteristics of the conjugates in applications will trigger the broader use of intramolecularPhotostabilization and help to emerge this approach as a new gold standard.
Abstract: Intramolecular photostabilization via triple-state quenching was recently revived as a tool to impart synthetic organic fluorophores with 'self-healing' properties. To date, utilization of such fluorophore derivatives is rare due to their elaborate multi-step synthesis. Here we present a general strategy to covalently link a synthetic organic fluorophore simultaneously to a photostabilizer and biomolecular target via unnatural amino acids. The modular approach uses commercially available starting materials and simple chemical transformations. The resulting photostabilizer-dye conjugates are based on rhodamines, carbopyronines and cyanines with excellent photophysical properties, that is, high photostability and minimal signal fluctuations. Their versatile use is demonstrated by single-step labelling of DNA, antibodies and proteins, as well as applications in single-molecule and super-resolution fluorescence microscopy. We are convinced that the presented scaffolding strategy and the improved characteristics of the conjugates in applications will trigger the broader use of intramolecular photostabilization and help to emerge this approach as a new gold standard.

90 citations

Journal ArticleDOI
TL;DR: In this article, a general synthetic route to carbopyronines with functional groups variable in the final steps of the synthesis or in the resulting fluorescent dye is presented, and a multistep synthesis of the dye starting from a simple precursor and utilizing a single temporary protective group is described.

88 citations

Journal ArticleDOI
TL;DR: In this article, hydrogenated rhodamines, carbopyronines, carbo-, silico-, and germanorhodamines with absorption maxima in the range of 530-640 µm were used for specific labeling of living cells.
Abstract: Hydroxylated rhodamines, carbopyronines, silico- and germanorhodamines with absorption maxima in the range of 530-640 nm were prepared and applied in specific labeling of living cells. The direct and high-yielding entry to germa- and silaxanthones tolerates the presence of protected heteroatoms and may be considered for the syntheses of various sila- and germafluoresceins, as well as -rhodols. Application in stimulated emission depletion (STED) fluorescence microscopy revealed a resolution of 50-75 nm in one- and two-color imaging of vimentin-HaloTag fused protein and native tubulin. The established structure-property relationships allow for prediction of the spectral properties and the positions of spirolactone/zwitterion equilibria for the new analogues of rhodamines, carbo-, silico-, and germanorhodamines using simple additive schemes.

73 citations


Cited by
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Journal ArticleDOI
TL;DR: The review covers the knowledge on photoremovable protecting groups and includes all relevant chromophores studied in the time period of 2000–2012 and the most relevant earlier works are discussed.
Abstract: The review covers the knowledge on photoremovable protecting groups and includes all relevant chromophores studied in the time period of 2000–2012; the most relevant earlier works are also discussed.

1,274 citations

Journal ArticleDOI
TL;DR: This review focuses on classifying different types of long wavelength absorbing BODIPY dyes based on the wide range of structural modification methods that have been adopted, and on tabulating their spectral and photophysical properties.
Abstract: This review focuses on classifying different types of long wavelength absorbing BODIPY dyes based on the wide range of structural modification methods that have been adopted, and on tabulating their spectral and photophysical properties. The structure–property relationships are analyzed in depth with reference to molecular modeling calculations, so that the effectiveness of the different structural modification strategies for shifting the main BODIPY spectral bands to longer wavelengths can be readily compared, along with their effects on the fluorescence quantum yield (ΦF) values. This should facilitate the future rational design of red/NIR region BODIPY dyes for a wide range of different applications.

1,013 citations

Journal ArticleDOI
TL;DR: A biocompatible near-infrared silicon-rhodamine probe that can be coupled specifically to proteins using different labelling techniques, and its high permeability and fluorogenic character permit the imaging of proteins in living cells and tissues.
Abstract: The ideal fluorescent probe for bioimaging is bright, absorbs at long wavelengths and can be implemented flexibly in living cells and in vivo. However, the design of synthetic fluorophores that combine all of these properties has proved to be extremely difficult. Here, we introduce a biocompatible near-infrared silicon-rhodamine probe that can be coupled specifically to proteins using different labelling techniques. Importantly, its high permeability and fluorogenic character permit the imaging of proteins in living cells and tissues, and its brightness and photostability make it ideally suited for live-cell super-resolution microscopy. The excellent spectroscopic properties of the probe combined with its ease of use in live-cell applications make it a powerful new tool for bioimaging.

740 citations

Journal ArticleDOI
TL;DR: It is reported that the nucleoids labeled with antibodies against DNA, mitochondrial transcription factor A (TFAM), or incorporated BrdU, have a defined, uniform mean size of ∼100 nm in mammals, and it is shown that TFAM is a main constituent of the nucleoid, besides mtDNA.
Abstract: Mammalian mtDNA is packaged in DNA-protein complexes denoted mitochondrial nucleoids. The organization of the nucleoid is a very fundamental question in mitochondrial biology and will determine tissue segregation and transmission of mtDNA. We have used a combination of stimulated emission depletion microscopy, enabling a resolution well below the diffraction barrier, and molecular biology to study nucleoids in a panel of mammalian tissue culture cells. We report that the nucleoids labeled with antibodies against DNA, mitochondrial transcription factor A (TFAM), or incorporated BrdU, have a defined, uniform mean size of ∼100 nm in mammals. Interestingly, the nucleoid frequently contains only a single copy of mtDNA (average ∼1.4 mtDNA molecules per nucleoid). Furthermore, we show by molecular modeling and volume calculations that TFAM is a main constituent of the nucleoid, besides mtDNA. These fundamental insights into the organization of mtDNA have broad implications for understanding mitochondrial dysfunction in disease and aging.

474 citations

Journal ArticleDOI
TL;DR: The future of this method in regard to spatiotemporal limits, live-cell imaging and combination with spectroscopy is discussed, and advances in these areas may elevate STED microscopy to a standard method for imaging in the life sciences.
Abstract: Stimulated emission depletion (STED) microscopy provides subdiffraction resolution while preserving useful aspects of fluorescence microscopy, such as optical sectioning, and molecular specificity and sensitivity However, sophisticated microscopy architectures and high illumination intensities have limited STED microscopy's widespread use in the past Here we summarize the progress that is mitigating these problems and giving substantial momentum to STED microscopy applications We discuss the future of this method in regard to spatiotemporal limits, live-cell imaging and combination with spectroscopy Advances in these areas may elevate STED microscopy to a standard method for imaging in the life sciences

393 citations