Leutenegger, Marcel

[["dc.bibliographiccitation.artnumber","7368"],["dc.bibliographiccitation.firstpage","204"],["dc.bibliographiccitation.journal","Nature"],["dc.bibliographiccitation.lastpage","208"],["dc.bibliographiccitation.volume","478"],["dc.contributor.author","Grotjohann, Tim"],["dc.contributor.author","Testa, Ilaria"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Bock, Hannes"],["dc.contributor.author","Urban, Nicolai T."],["dc.contributor.author","Lavoie-Cardinal, Flavie"],["dc.contributor.author","Willig, Katrin I."],["dc.contributor.author","Eggeling, Christian"],["dc.contributor.author","Jakobs, Stefan"],["dc.contributor.author","Hell, Stefan W."],["dc.date.accessioned","2017-09-07T11:43:21Z"],["dc.date.available","2017-09-07T11:43:21Z"],["dc.date.issued","2011"],["dc.description.abstract","Lens-based optical microscopy failed to discern fluorescent features closer than 200 nm for decades, but the recent breaking of the diffraction resolution barrier by sequentially switching the fluorescence capability of adjacent features on and off is making nanoscale imaging routine. Reported fluorescence nanoscopy variants switch these features either with intense beams at defined positions or randomly, molecule by molecule. Here we demonstrate an optical nanoscopy that records raw data images from living cells and tissues with low levels of light. This advance has been facilitated by the generation of reversibly switchable enhanced green fluorescent protein (rsEGFP), a fluorescent protein that can be reversibly photoswitched more than a thousand times. Distributions of functional rsEGFP-fusion proteins in living bacteria and mammalian cells are imaged at <40-nanometre resolution. Dendritic spines in living brain slices are super-resolved with about a million times lower light intensities than before. The reversible switching also enables all-optical writing of features with subdiffraction size and spacings, which can be used for data storage."],["dc.identifier.doi","10.1038/nature10497"],["dc.identifier.gro","3142644"],["dc.identifier.isi","000295782800041"],["dc.identifier.pmid","21909116"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/71"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","0028-0836"],["dc.title","Diffraction-unlimited all-optical imaging and writing with a photochromic GFP"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","26417"],["dc.bibliographiccitation.issue","25"],["dc.bibliographiccitation.journal","Optics Express"],["dc.bibliographiccitation.lastpage","26429"],["dc.bibliographiccitation.volume","18"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Eggeling, Christian"],["dc.contributor.author","Hell, Stefan"],["dc.date.accessioned","2017-09-07T11:45:09Z"],["dc.date.available","2017-09-07T11:45:09Z"],["dc.date.issued","2010"],["dc.description.abstract","Stimulated emission depletion (STED) resolves fluorescent features that are closer than the far-field optical diffraction limit by applying a spatially modulated light field keeping all but one of these features dark consecutively. For estimating the efficiency of transient fluorophore darkening, we developed analytical equations considering the spatio-temporal intensity profile of the STED beam. These equations provide a quick analysis and optimization of the resolution and contrast to be gained under various conditions, such as continuous wave or pulsed STED beams having different pulse durations. Particular emphasis is placed on fluorescence fluctuation methods such as correlation spectroscopy (FCS) using STED."],["dc.identifier.doi","10.1364/OE.18.026417"],["dc.identifier.gro","3142820"],["dc.identifier.isi","000285749500100"],["dc.identifier.pmid","21164992"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/266"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","1094-4087"],["dc.title","Analytical description of STED microscopy performance"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.artnumber","040502"],["dc.bibliographiccitation.issue","4"],["dc.bibliographiccitation.journal","Journal of Biomedical Optics"],["dc.bibliographiccitation.volume","11"],["dc.contributor.author","Leutenegger, M."],["dc.contributor.author","Blom, H."],["dc.contributor.author","Widengren, J."],["dc.contributor.author","Eggeling, C."],["dc.contributor.author","Goesch, M."],["dc.contributor.author","Leitgeb, R. A."],["dc.contributor.author","Lasser, T."],["dc.date.accessioned","2018-03-19T22:46:29Z"],["dc.date.available","2018-03-19T22:46:29Z"],["dc.date.issued","2006"],["dc.description.abstract","We present the development and first application of a novel dual-color total internal reflection (TIR) fluorescence system for single-molecule coincidence analysis and fluorescence cross-correlation spectroscopy (FCCS). As a performance analysis, we measured a synthetic DNA-binding assay, demonstrating this dual-color TIR-FCCS approach to be a suitable method for measuring coincidence assays such as biochemical binding, fusion, or signal transduction at solid/liquid interfaces. Due to the very high numerical aperture of the epi-illumination configuration, our setup provides a very high fluorescence collection efficiency resulting in a two- to three-fold increase in molecular brightness compared to conventional confocal FCCS. Further improvements have been achieved through global analysis of the spectroscopic data."],["dc.identifier.doi","10.1117/1.2221714"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/13090"],["dc.language.iso","en"],["dc.notes.status","final"],["dc.title","Dual-color total internal reflection fluorescence cross-correlation spectroscopy"],["dc.type","journal_article"],["dc.type.internalPublication","unknown"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","771"],["dc.bibliographiccitation.issue","4"],["dc.bibliographiccitation.journal","ChemPhysChem"],["dc.bibliographiccitation.lastpage","783"],["dc.bibliographiccitation.volume","15"],["dc.bibliographiccitation.volumetitle","Superresolution Imaging and Nanophotonics"],["dc.contributor.author","Sahl, Steffen J."],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Hell, Stefan"],["dc.contributor.author","Eggeling, Christian"],["dc.date.accessioned","2017-09-07T11:46:25Z"],["dc.date.available","2017-09-07T11:46:25Z"],["dc.date.issued","2014"],["dc.description.abstract","The performance of a method is assessed which allows for the spatiotemporal tracking of single dye-labeled molecules during two-dimensional (2D) diffusional transits through the focal area of a modified confocal microscope. In addition to facilitating the observation of molecular diffusion paths at the shot-noise limit of bright organic emitters with spatial and temporal precisions of approximate to 10-20 nm and <0.5 ms, respectively, the direct access to the complete stream of detected photons is beneficial for characterizing nanoscale details such as transient pausing (binding). We discuss technical aspects of this approach, along with results from its application to measuring lipid membrane dynamics in live mammalian cells. Presented topics include a discussion of the advantages of the single-photon collection mode and instrument as well as computational considerations for the localization process. A proof-of-principle experiment shows that optical nanoscopy by stochastic single-molecule switching and position readout could be implementable in parallel with such fast molecular tracking. This would allow direct access to contextual imaging data of local cytoskeletal structural elements or localized longer-lived protein assemblies."],["dc.identifier.doi","10.1002/cphc.201301090"],["dc.identifier.gro","3142170"],["dc.identifier.isi","000332747500028"],["dc.identifier.pmid","24596277"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/5310"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10 / Funder: Deutsche Forschungsgemeinschaft; DFG [SFB755]"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.eissn","1439-7641"],["dc.relation.issn","1439-4235"],["dc.title","High-Resolution Tracking of Single-Molecule Diffusion in Membranes by Confocalized and Spatially Differentiated Fluorescence Photon Stream Recording"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","3593"],["dc.bibliographiccitation.issue","19"],["dc.bibliographiccitation.journal","European Journal of Organic Chemistry"],["dc.bibliographiccitation.lastpage","3610"],["dc.bibliographiccitation.volume","2010"],["dc.contributor.author","Kolmakov, Kirill"],["dc.contributor.author","Belov, Vladimir N."],["dc.contributor.author","Wurm, Christian Andreas"],["dc.contributor.author","Harke, Benjamin"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Eggeling, Christian"],["dc.contributor.author","Hell, Stefan"],["dc.date.accessioned","2017-09-07T11:45:57Z"],["dc.date.available","2017-09-07T11:45:57Z"],["dc.date.issued","2010"],["dc.description.abstract","Biological microscopy favors photostable fluorescent markers with large fluorescence quantum yields, low dark triplet state population, good biocompatibility and absorption and emission maxima in the near-infrared, where cellular autofluorescence is minimized. In the present study, carbopyronines absorbing around 640 nm and emitting at around 660 nm, with a low intersystem crossing rate (k(isc) approximate to 0.5 X 10(6) s(-1)) and excellent properties for cellular imaging were synthesized. 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. Possessing two 2-methoxyethyl groups, the parent dye is soluble in water and most organic solvents. Demethylation of the dye or its precursors is straightforward, clean, and furnishes compounds with one or two 2-hydroxyethyl groups, which can be used for further transformations. Modifications in the linker-containing carboxy group are also possible. A multistep synthesis of the dye starting from a simple precursor and utilizing a single temporary protective group is described. The presented approach may be further applied to the design of caged carbopyronines."],["dc.identifier.doi","10.1002/ejoc.201000343"],["dc.identifier.gro","3142895"],["dc.identifier.isi","000280220800007"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/350"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10 / Funder: Deutsche Forschungsgemeinschaft (DFG); Max-Planck-Gesellschaft"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","1434-193X"],["dc.title","A Versatile Route to Red-Emitting Carbopyronine Dyes for Optical Microscopy and Nanoscopy"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","5243"],["dc.bibliographiccitation.issue","5"],["dc.bibliographiccitation.journal","Optics Express"],["dc.bibliographiccitation.lastpage","5263"],["dc.bibliographiccitation.volume","20"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Ringemann, Christian"],["dc.contributor.author","Lasser, Theo"],["dc.contributor.author","Hell, Stefan"],["dc.contributor.author","Eggeling, Christian"],["dc.date.accessioned","2017-09-07T11:48:59Z"],["dc.date.available","2017-09-07T11:48:59Z"],["dc.date.issued","2012"],["dc.description.abstract","We characterize a novel fluorescence microscope which combines the high spatial discrimination of a total internal reflection epi-fluorescence (epi-TIRF) microscope with that of stimulated emission depletion (STED) nanoscopy. This combination of high axial confinement and dynamic-active lateral spatial discrimination of the detected fluorescence emission promises imaging and spectroscopy of the structure and function of cell membranes at the macro-molecular scale. Following a full theoretical description of the sampling volume and the recording of images of fluorescent beads, we exemplify the performance and limitations of the TIRF-STED nanoscope with particular attention to the polarization state of the laser excitation light. We demonstrate fluorescence correlation spectroscopy (FCS) with the TIRF-STED nanoscope by observing the diffusion of dye molecules in aqueous solutions and of fluorescent lipid analogs in supported lipid bilayers in the presence of background signal. The nanoscope reduced the out-of-focus background signal. A lateral resolution down to 40-50 nm was attained which was ultimately limited by the low lateral signal-to-background ratio inherent to the confocal epi-TIRF scheme. Together with the estimated axial confinement of about 55 nm, our TIRF-STED nanoscope achieved an almost isotropic and less than 1 attoliter small all-optically induced measurement volume."],["dc.identifier.doi","10.1364/OE.20.005243"],["dc.identifier.gro","3142578"],["dc.identifier.isi","000301053200045"],["dc.identifier.pmid","22418331"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/8944"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","1094-4087"],["dc.title","Fluorescence correlation spectroscopy with a total internal reflection fluorescence STED microscope (TIRF-STED-FCS)"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.journal","Protein Science"],["dc.bibliographiccitation.volume","21"],["dc.contributor.author","Grotjohann, Tim"],["dc.contributor.author","Testa, Ilaria"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Eggeling, Christian"],["dc.contributor.author","Hell, Stefan W."],["dc.contributor.author","Jakobs, Stefan"],["dc.date.accessioned","2018-11-07T09:07:54Z"],["dc.date.available","2018-11-07T09:07:54Z"],["dc.date.issued","2012"],["dc.format.extent","163"],["dc.identifier.isi","000307019800279"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/25904"],["dc.notes.status","zu prüfen"],["dc.notes.submitter","Najko"],["dc.publisher","Wiley-blackwell"],["dc.publisher.place","Hoboken"],["dc.relation.eventlocation","San Diego, CA"],["dc.relation.issn","0961-8368"],["dc.title","Reversibly switchable Enhanced Green Fluorescent Protein (rsEGFP): a novel photochromic fluorescent protein enables RESOLFT-type microscopy of living cells at low light intensities"],["dc.type","conference_abstract"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.status","published"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","942"],["dc.bibliographiccitation.issue","10"],["dc.bibliographiccitation.journal","Nature Biotechnology"],["dc.bibliographiccitation.lastpage","U132"],["dc.bibliographiccitation.volume","29"],["dc.contributor.author","Brakemann, Tanja"],["dc.contributor.author","Stiel, Andre C."],["dc.contributor.author","Weber, Gert"],["dc.contributor.author","Andresen, Martin"],["dc.contributor.author","Testa, Ilaria"],["dc.contributor.author","Grotjohann, Tim"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Plessmann, Uwe"],["dc.contributor.author","Urlaub, Henning"],["dc.contributor.author","Eggeling, Christian"],["dc.contributor.author","Wahl, Markus C."],["dc.contributor.author","Hell, Stefan"],["dc.contributor.author","Jakobs, Stefan"],["dc.date.accessioned","2017-09-07T11:43:22Z"],["dc.date.available","2017-09-07T11:43:22Z"],["dc.date.issued","2011"],["dc.description.abstract","Photoswitchable fluorescent proteins have enabled new approaches for imaging cells, but their utility has been limited either because they cannot be switched repeatedly or because the wavelengths for switching and fluorescence imaging are strictly coupled. We report a bright, monomeric, reversibly photoswitchable variant of GFP, Dreiklang, whose fluorescence excitation spectrum is decoupled from that for optical switching. Reversible on-and-off switching in living cells is accomplished at illumination wavelengths of similar to 365 nm and similar to 405 nm, respectively, whereas fluorescence is elicited at similar to 515 nm. Mass spectrometry and high-resolution crystallographic analysis of the same protein crystal in the photoswitched on- and off-states demonstrate that switching is based on a reversible hydration/dehydration reaction that modifies the chromophore. The switching properties of Dreiklang enable far-field fluorescence nanoscopy in living mammalian cells using both a coordinate-targeted and a stochastic single molecule switching approach."],["dc.identifier.doi","10.1038/nbt.1952"],["dc.identifier.gro","3142656"],["dc.identifier.isi","000296273000022"],["dc.identifier.pmid","21909082"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/84"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.publisher","Nature Publishing Group"],["dc.relation.eissn","1546-1696"],["dc.relation.issn","1087-0156"],["dc.title","A reversibly photoswitchable GFP-like protein with fluorescence excitation decoupled from switching"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.artnumber","103054"],["dc.bibliographiccitation.journal","New Journal of Physics"],["dc.bibliographiccitation.volume","11"],["dc.contributor.author","Ringemann, Christian"],["dc.contributor.author","Harke, Benjamin"],["dc.contributor.author","von Middendorff, Claas"],["dc.contributor.author","Medda, Rebecca"],["dc.contributor.author","Honigmann, Alf"],["dc.contributor.author","Wagner, Richard"],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Schönle, Andreas"],["dc.contributor.author","Hell, Stefan"],["dc.contributor.author","Eggeling, Christian"],["dc.date.accessioned","2017-09-07T11:46:48Z"],["dc.date.available","2017-09-07T11:46:48Z"],["dc.date.issued","2009"],["dc.description.abstract","The study of molecular dynamics at the single-molecule level with fluorescence correlation spectroscopy (FCS) and far-field optics has contributed greatly to the functional understanding of complex systems. Unfortunately, such studies are restricted to length scales of >200 nm because diffraction does not allow further reduction of the measurement volume. This sets an upper limit on the applicable concentration of fluorescently labeled molecules and even more importantly, averages out details of nanoscale dynamics. By combining FCS and fluorescence intensity distribution analysis (FIDA) with sub-diffraction-resolution stimulated emission depletion (STED) nanoscopy, we remove this restriction and obtain open measurement volumes of nanoscale dimensions which are tunable in size. As a consequence, single-molecule studies can now be extended to nanoscale dynamics and may be applied to much larger, often endogenous concentrations. In solution, low-brightness signal from axial out-of-focus volume shells was taken into account by using both FCS and FIDA in conjunction to analyze the data. In two-dimensional systems, such as lipid membranes, the background is greatly reduced and measurements feature excellent signal-to-noise ratios. Measurement foci of down to 30 nm in diameter directly reveal anomalous diffusion of lipids in the plasma membrane of living cells and allow for the determination of on) off rates of the binding of lipids to other membrane constituents. Such important insight into the prominent biological question of lipid membrane organization or 'lipid rafts' shows that combining fluctuation analysis with STED-engineered ultra-small measurement volumes is a viable and powerful new approach to probing molecular dynamics on the nanoscale."],["dc.identifier.doi","10.1088/1367-2630/11/10/103054"],["dc.identifier.gro","3143036"],["dc.identifier.isi","000271324700005"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/506"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","1367-2630"],["dc.title","Exploring single-molecule dynamics with fluorescence nanoscopy"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]

[["dc.bibliographiccitation.firstpage","6829"],["dc.bibliographiccitation.issue","15"],["dc.bibliographiccitation.journal","Proceedings of the National Academy of Sciences"],["dc.bibliographiccitation.lastpage","6834"],["dc.bibliographiccitation.volume","107"],["dc.contributor.author","Sahl, Steffen J."],["dc.contributor.author","Leutenegger, Marcel"],["dc.contributor.author","Hilbert, Michael"],["dc.contributor.author","Hell, Stefan"],["dc.contributor.author","Eggeling, Christian"],["dc.date.accessioned","2017-09-07T11:46:06Z"],["dc.date.available","2017-09-07T11:46:06Z"],["dc.date.issued","2010"],["dc.description.abstract","We describe an optical method capable of tracking a single fluorescent molecule with a flexible choice of high spatial accuracy (similar to 10-20 nm standard deviation or similar to 20-40 nm full-width-at-half-maximum) and temporal resolution (<1 ms). The fluorescence signal during individual passages of fluorescent molecules through a spot of excitation light allows the sequential localization and thus spatio-temporal tracking of the molecule if its fluorescence is collected on at least three separate point detectors arranged in close proximity. We show two-dimensional trajectories of individual, small organic dye labeled lipids diffusing in the plasma membrane of living cells and directly observe transient events of trapping on <20 nm spatial scales. The trapping is cholesterol-assisted and much more pronounced for a sphingo- than for a phosphoglycero-lipid, with average trapping times of similar to 15 ms and <4 ms, respectively. The results support previous STED nanoscopy measurements and suggest that, at least for nontreated cells, the transient interaction of a single lipid is confined to macromolecular dimensions. Our experimental approach demonstrates that fast molecular movements can be tracked with minimal invasion, which can reveal new important details of cellular nano-organization."],["dc.identifier.doi","10.1073/pnas.0912894107"],["dc.identifier.gro","3142937"],["dc.identifier.isi","000276642100050"],["dc.identifier.pmid","20351247"],["dc.identifier.uri","https://resolver.sub.uni-goettingen.de/purl?gro-2/396"],["dc.language.iso","en"],["dc.notes.intern","WoS Import 2017-03-10 / Funder: Leibniz-Prize"],["dc.notes.status","final"],["dc.notes.submitter","PUB_WoS_Import"],["dc.relation.issn","0027-8424"],["dc.title","Fast molecular tracking maps nanoscale dynamics of plasma membrane lipids"],["dc.type","journal_article"],["dc.type.internalPublication","yes"],["dc.type.peerReviewed","yes"],["dc.type.subtype","original"],["dspace.entity.type","Publication"]]