Dokument-ID Dokumenttyp Autoren/innen Persönliche Herausgeber/innen Haupttitel Abstract Auflage Verlagsort Verlag Herausgeber (Institution) Erscheinungsjahr Titel des übergeordneten Werkes Jahrgang/Band ISBN Veranstaltung Veranstaltungsort Beginndatum der Veranstaltung Enddatum der Veranstaltung Ausgabe/Heft Erste Seite Letzte Seite URN DOI Lizenz Datum der Freischaltung OPUS4-31629 Zeitschriftenartikel Rades, Steffi; Hodoroaba, Vasile-Dan; Salge, T.; Wirth, Thomas; Lobera, M.P.; Labrador, R.H.; Natte, Kishore; Behnke, Thomas; Gross, Thomas; Unger, Wolfgang High-resolution imaging with SEM/T-SEM, EDX and SAM as a combined methodical approach for morphological and elemental analyses of single engineered nanoparticles The combination of complementary characterization techniques such as SEM (Scanning Electron Microscopy), T-SEM (Scanning Electron Microscopy in Transmission Mode), EDX (Energy Dispersive X-ray Spectroscopy) and SAM (Scanning Auger Microscopy) has been proven to be a powerful and relatively quick characterization strategy for comprehensive morphological and chemical characterization of individual silica and titania nanoparticles. The selected “real life” test materials, silica and titania, are listed in the OECD guidance manual as representative examples because they are often used as commercial nanomaterials. Imaging by high resolution SEM and in the transmission mode by T-SEM allows almost simultaneous surface and in-depth inspection of the same particle using the same instrument. EDX and SAM enable the chemical characterization of bulk and surface of individual nanoparticles. The core-shell properties of silica based materials are addressed as well. Titania nominally coated by silane purchased from an industrial source has been found to be inhomogeneous in terms of chemical composition. London RSC Publishing 2014 RSC Advances 4 91 49577 49587 urn:nbn:de:kobv:b43-316296 10.1039/c4ra05092d http://creativecommons.org/licenses/by/3.0/de/deed.de 2016-02-20 OPUS4-37561 Zeitschriftenartikel Moser, Marko; Schneider, Ralf; Behnke, Thomas; Schneider, Thomas; Falkenhagen, Jana; Resch-Genger, Ute Ellman's and aldrithiol assay as versatile and complementary tools for the quantification of thiol groups and ligands on nanomaterials Simple, fast, and versatile methods for the quantification of thiol groups are of considerable interest not only for protein analysis but also for the characterization of the surface chemistry of nanomaterials stabilized with thiol ligands or bearing thiol groups for the subsequent (bio-) functionalization via maleimide−thiol chemistry. Here, we compare two simple colorimetric assays, the widely used Ellman's assay performed at alkaline pH and the aldrithiol assay executed at acidic and neutral pH, with respect to their potential for the quantification of thiol groups and thiol ligands on different types of nanoparticles like polystyrene nanoparticles, semiconductor nanocrystals (SC NC), and noble metal particles, and we derive criteria for their use. In order to assess the underlying reaction mechanisms and to obtain stoichiometry factors mandatory for reliable thiol quantification, both methods were studied photometrically and with electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS), thereby demonstrating the influence of different thiols on the reaction mechanism. Our results underline the suitability of both methods for the quantification of directly accessible thiol groups or ligands on the surface of 2D- and 3D-supports, here exemplarily polystyrene nanoparticles. Moreover, we could derive strategies for the use of these simple assays for the determination of masked (i.e., not directly accessible) thiol groups like disulfides such as lipoic acid and thiol stabilizing ligands coordinatively bound to Cd and/or Hg surface atoms of II/VI and ternary SC NC and to gold and silver nanoparticles. Washington ACS Publications 2016 Analytical Chemistry 88 17 8624 8631 10.1021/acs.analchem.6b01798 2016-09-29 OPUS4-45176 Zeitschriftenartikel Nirmalananthan-Budau, Nithiya; Behnke, Thomas; Hoffmann, Katrin; Kage, Daniel; Gers-Panther, Charlotte F.; Frank, Walter; Müller, Thomas J. J.; Resch-Genger, Ute Crystallization and Aggregation-Induced Emission in a Series of Pyrrolidinylvinylquinoxaline Derivatives Aggregation-induced emission (AIE) has been meanwhile observed for many dye classes and particularly for fluorophores containing propeller-like groups. Herein, we report on the AIE characteristics of a series of four hydrophobic pyrrolidinylvinylquinoxaline (PVQ) derivatives with phenyl, pyrrolyl, indolyl, and methoxythienyl substituents used to systematically vary the torsion angle between this substituent at the quinoxaline C2 position and the planar PVQ moiety. These molecules, which are accessible via four- or five-component one-pot syntheses, were spectroscopically studied in organic solvents and solvent−water mixtures, as dye aggregates, solids, and entrapped in polystyrene particles (PSP). Steady-state and time-resolved fluorescence measurements revealed a strong fluorescence enhancement for all dyes in ethanol−water mixtures of high water content, accompanying the formation of dye aggregates with sizes of a few hundred nm, overcoming polarity and H-bonding-induced fluorescence quenching of the chargetransfer-type emission of these PVQ dyes. The size and shape of these dye aggregates and the size of the AIE effect are controlled by the water content and the substituent-dependent torsion angle that influences the nucleation process and the packing of the molecules during aggregation. Staining of 1 μm-sized carboxy-functionalized PSP with the PVQ dyes resulted also in a considerable increase in the fluorescence quantum yield and lifetime, reflecting the combined influence of the restricted molecular motion and the reduced polarity of the dye microenvironment. ACS Publications 2018 Journal of Physical Chemistry c 122 20 11119 11127 10.1021/acs.jpcc.8b01425 2018-06-14 OPUS4-28466 Posterpräsentation Behnke, Thomas Design and Characterization of NIR-emissive PEGylated and Protein-functionalized Nanoparticles for Bioanalytics 2013 7th Senftenberger Innovationsforum Multiarameteranalytik 7th Senftenberger Innovationsforum Multiarameteranalytik Senftenberg, Germany 2013-04-18 2013-04-19 2016-02-20 OPUS4-28468 Posterpräsentation Behnke, Thomas Targeted NIR-emissive nanosensors for bioimaging 2013 7th Senftenberger Innovationsforum Multiparameteranalytik 7th Senftenberger Innovationsforum Multiparameteranalytik Senftenberg, Germany 2013-04-18 2013-04-19 2016-02-20 OPUS4-25698 Vortrag Behnke, Thomas Design and Bioanalytical Applications of NIR-Fluorescent Polystyrene Nanoparticles 2012 Imaging Innovations of the Lung Imaging Innovations of the Lung Göttingen, Germany 2012-04-18 2012-04-20 2016-02-19 OPUS4-26020 Posterpräsentation Behnke, Thomas Targeted ratiometric Nanosensors for the in Vivo Imaging of Tumor Hypoxia in the NIR spectral region 2012 Tag der Chemie Tag der Chemie Regensburg, Germany 2012-06-28 2016-02-19 OPUS4-26021 Posterpräsentation Behnke, Thomas Rational Design and Absolute Quantum yields of NIR-Fluorescent Nanoparticls for Bioanalytical Applications 2012 Innovationsforum "Funktionelle Farbstoffe" Innovationsforum "Funktionelle Farbstoffe" Bitterfeld-Wolfen, Germany 2012-05-31 2012-06-01 2016-02-19 OPUS4-26022 Posterpräsentation Behnke, Thomas NIR-Fluorescent Polystyrene Nanoparticles: Design, Optical Characterization and Bioanalytical Applications 2012 Abteilungsseminar BfR Abteilungsseminar BfR Berlin, Germany 2012-06-15 2016-02-19 OPUS4-24353 Beitrag zu einem Tagungsband Pauli, Jutta; Brehm, Robert; Grabolle, Markus; Behnke, Thomas; Mathejczyk, J.; Hamann, F.; Alves, F.; Hilger, I.; Resch-Genger, Ute Achilefu, S.; Raghavachari, R. Dye-biomolecule conjugates and NIR-fluorescent particles for targeting of disease-related biomarkers Indispensable for fluorescence imaging are highly specific and sensitive molecular probes that absorb and emit in the near infrared (NIR) spectral region and respond to or target molecular species or processes. Here, we present approaches to targeted fluorescent probes for in vivo imaging in the intensity and lifetime domain exploiting NIR dyes. Screening schemes for the fast identification of suitable fluorophores are derived and design criteria for highly emissive optical probes. In addition, as a signal amplification strategy that enables also the use of hydrophobic NIR fluorophores as fluorescent reporters, first steps towards versatile strategies for the preparation of NIR-fluorescent polymeric particles are presented that can be utilized also for the design of targeted and analyte-responsive probes. The international society for optics and photonics (SPIE) 2011 Proceedings of SPIE - Reporters, markers, dyes, nanoparticles, and molecular probes for biomedical applications III 7910 791014-1 791014-15 10.1117/12.876828 2016-02-19 OPUS4-24987 Zeitschriftenartikel Napp, J.; Behnke, Thomas; Fischer, L.; Würth, Christian; Wottawa, M.; Katschinski, D.M.; Alves, F.; Resch-Genger, Ute; Schäferling, M. Targeted luminescent near-infrared polymer-nanoprobes for in vivo imaging of tumor hypoxia Polystyrene nanoparticles (PS-NPs) were doped with an oxygen-sensitive near-infrared (NIR)-emissive palladium meso-tetraphenylporphyrin and an inert reference dye which are both excitable at 635 nm. The nanosensors were characterized with special emphasis on fundamental parameters such as absolute photoluminescence quantum yield and fluorescence lifetime. The PS-NPs were employed for ratiometric dual-wavelength and lifetime-based photoluminescent oxygen sensing. They were efficiently taken up by cultured murine alveolar macrophages, yielding a characteristic and reversible change in ratiometric response with decreasing oxygen concentration. This correlated with the cellular hypoxic status verified by analysis of hypoxia inducible factor-1α (HIF-1α) accumulation. In addition, the surface of PS-NPs was functionalized with polyethylene glycol (PEG) and the monoclonal antibody herceptin, and their binding to HER2/neu-overexpressing tumor cells was confirmed in vitro. First experiments with tumor-bearing mouse revealed a distinctive ratiometric response within the tumor upon hypoxic condition induced by animal sacrifice. These results demonstrate the potential of these referenced NIR nanosensors for in vitro and in vivo imaging that present a new generation of optical probes for oncology. Washington, DC American Chemical Society 2011 Analytical chemistry 83 23 9039 9046 10.1021/ac201870b 2016-02-19 OPUS4-27682 Zeitschriftenartikel Behnke, Thomas; Laux, Eva-Maria; Hoffmann, Katrin; Peters, S.; Haueisen, J.; Klemm, M.; Resch-Genger, Ute Simple approaches to fluorescence lifetime standards using dye-quencher pairs Photoluminescence techniques are amongst the most widely used tools in the material and life sciences, with new and exciting applications continuously emerging. Advantages include their comparative ease of use, unique sensitivity, non-invasive character, and potential for multiplexing, remote sensing, and miniaturization. Increasing applications of fluorescence techniques in the life sciences and emerging medical applications of fluorescence microscopic techniques including 1P and 2P fluorescence microscopy combined with fluorescence lifetime imaging (FLIM) in e.g. in vivo eye diagnostics boosted the demand for robust, easy-to-use, and reliable fluorescence standards to ensure the reliability and comparability of fluorescence data. This includes fluorescence standards for the consideration of instrument-specific spectral and intensity distortions of measured signals and instrument performance validation, fluorescence intensity standards for the quantification of measured intensities and for signal referencing, and lately, also fluorescence lifetime standards. Berlin De Gruyter Österreichische Gesellschaft für Biomedizinische Technik 2012 Biomedizinische Technik = Biomedical engineering 57 BMT 2012 - 46. DGBMT Jahrestagung Jena, Germany 16.09.2012 19.09.2012 Suppl. 1 613 10.1515/bmt-2012-4499 2016-02-20 OPUS4-25872 Zeitschriftenartikel Natte, Kishore; Behnke, Thomas; Orts Gil, Guillermo; Würth, Christian; Friedrich, Jörg Florian; Österle, Werner; Resch-Genger, Ute Synthesis and characterisation of highly fluorescent core-shell nanoparticles based on Alexa dyes Current and future developments in the emerging field of nanobiotechnology are closely linked to the rational design of novel fluorescent nanomaterials, e.g. for biosensing and imaging applications. Here, the synthesis of bright near infrared (NIR)-emissive nanoparticles based on the grafting of silica nanoparticles (SNPs) with 3-aminopropyl triethoxysilane (APTES) followed by covalent attachment of Alexa dyes and their subsequent shielding by an additional silica shell are presented. These nanoparticles were investigated by dynamic light scattering (DLS), transmission electron microscopy (TEM) and fluorescence spectroscopy. TEM studies revealed the monodispersity of the initially prepared and fluorophorelabelled silica particles and the subsequent formation of raspberry-like structures after addition of a silica precursor.Measurements of absolute fluorescence quantum yields of these scattering particle suspensions with an integrating sphere setup demonstrated the influence of dye labelling density-dependent fluorophore aggregation on the signaling behaviour of such nanoparticles. Dordrecht Kluwer 2012 Journal of nanoparticle research 14 2 680-1 - 680-10 10.1007/s11051-011-0680-9 2016-02-19 OPUS4-25873 Zeitschriftenartikel Huber, Alexandra; Behnke, Thomas; Würth, Christian; Jäger, Christian; Resch-Genger, Ute Spectroscopic characterization of coumarin-stained beads: quantification of the number of fluorophores per particle with solid-state 19F-NMR and measurement of absolute fluorescence quantum yields The rational design of nano- and micrometer-sized particles with tailor-made optical properties for biological, diagnostic, and photonic applications requires tools to characterize the signal-relevant properties of these typically scattering bead suspensions. This includes methods for the preferably nondestructive quantification of the number of fluorophores per particle and the measurement of absolute fluorescence quantum yields and absorption coefficients of suspensions of fluorescent beads for material performance optimization and comparison. Here, as a first proof-of-concept, we present the first time determination of the number of dye molecules per bead using nondestructive quantitative (19F) NMR spectroscopy and 1000 nm-sized carboxylated polystyrene particles loaded with varying concentrations of the laser dye coumarin 153 containing a CF3 group. Additionally, the signal-relevant optical properties of these dye-loaded particles were determined in aqueous suspension in comparison to the free dye in solvents of different polarity with a custom-built integrating sphere setup that enables spectrally resolved measurements of emission, transmission, and reflectance as well absolute fluorescence quantum yields. These measurements present an important step toward absolute brightness values and quantitative fluorescence analysis with particle systems that can be exploited, for example, for optical imaging techniques and different fluorescence assays as well as for the metrological traceability of fluorescence methods. Washington, DC American Chemical Society 2012 Analytical chemistry 84 8 3654 3661 10.1021/ac3000682 2016-02-19 OPUS4-27808 Posterpräsentation Behnke, Thomas Bioimaging with rationally designed NIR-flourescent nanosensors 2013 NAMDIATREAM Winter School on Bioimaging, sensing and therapeutic applications of nanomaterials NAMDIATREAM Winter School on Bioimaging, sensing and therapeutic applications of nanomaterials Villars-sur-Ollon, Switzerland 2013-03-03 2013-03-06 2016-02-20 OPUS4-27810 Posterpräsentation Behnke, Thomas NIR-Emissive O2-Nanosensors for Biomarker Imaging 2013 Biosensor Symposium Biosensor Symposium Wildau, Germany 2013-03-10 2013-03-13 2016-02-20 OPUS4-29017 Zeitschriftenartikel Felbeck, Tom; Behnke, Thomas; Hoffmann, Katrin; Grabolle, Markus; Lezhnina, M.M.; Kynast, U.H.; Resch-Genger, Ute Nile-red-nanoclay hybrids: Red emissive optical probes for use in aqueous dispersion Water-dispersible and (bio)functionalizable nanoclays have a considerable potential as inexpensive carriers for organic molecules like drugs and fluorophores. Aiming at simple design strategies for red-emissive optical probes for the life sciences from commercial precursors with minimum synthetic effort, we systematically studied the dye loading behavior and stability of differently functionalized laponites. Here, we present a comprehensive study of the absorption and emission properties of the red emissive hydrophobic and neutral dye Nile Red, a well-known polarity probe, which is almost insoluble and nonemissive in water. Adsorption of this probe onto disk-shaped nanoclays was studied in aqueous dispersion as function of dye concentration, in the absence and presence of the cationic surfactant cetyltrimethylammonium bromide (CTAB) assisting dye loading, and as a function of pH. This laponite loading strategy yields strongly fluorescent nanoclay suspensions with a fluorescence quantum yield of 0.34 at low dye loading concentration. The dye concentration-, CTAB-, and pH-dependent absorption, fluorescence emission, and fluorescence excitation spectra of the Nile-Red6#8211;nanoclay suspensions suggest the formation of several Nile Red species including emissive Nile Red monomers facing a polar environment, nonemissive H-type dimers, and protonated Nile Red molecules that are also nonfluorescent. Formation of all nonemissive Nile Red species could be suppressed by modification of the laponite with CTAB. This underlines the great potential of properly modified and functionalized laponite nanodisks as platform for optical probes with drug delivery capacities, for example, for tumor and therapy imaging. Moreover, comparison of the Nile Red dimer absorption spectra with absorption spectra of previously studied Nile Red aggregates in dendrimer systems and micelles and other literature systems reveals a considerable dependence of the dimer absorption band on microenvironment polarity which has not yet been reported so far for H-type dye aggregates. Washington, DC American Chemical Society 2013 Langmuir 29 36 11489 11497 10.1021/la402165q 2016-02-20 OPUS4-29031 Zeitschriftenartikel Hoffmann, Katrin; Behnke, Thomas; Drescher, Daniela; Kneipp, Janina; Resch-Genger, Ute Near-infrared-emitting nanoparticles for lifetime-based multiplexed analysis and imaging of living cells The increase in information content from bioassays and bioimaging requires robust and efficient strategies for the detection of multiple analytes or targets in a single measurement, thereby addressing current health and security concerns. For fluorescence techniques, an attractive alternative to commonly performed spectral or color multiplexing presents lifetime multiplexing and the discrimination between different fluorophores based on their fluorescence decay kinetics. This strategy relies on fluorescent labels with sufficiently different lifetimes that are excitable at the same wavelength and detectable within the same spectral window. Here, we report on lifetime multiplexing and discrimination with a set of nanometer-sized particles loaded with near-infrared emissive organic fluorophores chosen to display very similar absorption and emission spectra, yet different fluorescence decay kinetics in suspension. Furthermore, as a first proof-of-concept, we describe bioimaging studies with 3T3 fibroblasts and J774 macrophages, incubated with mixtures of these reporters employing fluorescence lifetime imaging microscopy. These proof-of-concept measurements underline the potential of fluorescent nanoparticle reporters in fluorescence lifetime multiplexing, barcoding, and imaging for cellular studies, cell-based assays, and molecular imaging. Washington, DC, USA ACS Publ. American Chemical Society 2013 ACS nano 7 8 6674 6684 10.1021/nn4029458 2016-02-20 OPUS4-22692 Zeitschriftenartikel Behnke, Thomas; Würth, Christian; Hoffmann, Katrin; Hübner, Martin; Panne, Ulrich; Resch-Genger, Ute Encapsulation of hydrophobic dyes in polystyrene micro- and nanoparticles via swelling procedures Aiming at the derivation of a generalized procedure for the straightforward preparation of particles fluorescing in the visible and near-infrared (NIR) spectral region, different swelling procedures for the loading of the hydrophobic polarity-probe Nile Red into nano- and micrometer sized polystyrene particles were studied and compared with respect to the optical properties of the resulting particles. The effect of the amount of incorporated dye on the spectroscopic properties of the particles was investigated for differently sized beads with different surface chemistries, i.e., non-functionalized, aminomodified and PEG-grafted surfaces. Moreover, photostability and leaking studies were performed. The main criterion for the optimization of the dye loading procedures was a high and thermally and photochemically stable fluorescence output of the particles for the future application of these systems as fluorescent labels. New York, NY Plenum Publ. Corp. 2011 Journal of fluorescence 21 3 937 944 10.1007/s10895-010-0632-2 2016-02-19 OPUS4-20642 Posterpräsentation Behnke, Thomas NIR-floureszente Mikro- und Nanopartikel für die Bioanalytik 2009 Zukunft Made in Adlershof Zukunft Made in Adlershof Berlin, Germany 2009-09-08 2016-02-19 OPUS4-22073 Posterpräsentation Behnke, Thomas NIR-Fluorescent Nano- and Micrometer-sized Particles for Biomedical and Diagnostical Applications 2010 Faraday Discussion, University of Edinburgh Faraday Discussion, University of Edinburgh Edinburgh, Scotland 2010-09-06 2010-09-08 2016-02-19 OPUS4-26877 Zeitschriftenartikel Behnke, Thomas; Mathejczyk, J.E.; Brehm, Robert; Würth, Christian; Gomes, F.R.; Dullin, C.; Napp, J.; Alves, F.; Resch-Genger, Ute Target-specific nanoparticles containing a broad band emissive NIR dye for the sensitive detection and characterization of tumor development Current optical probes including engineered nanoparticles (NPs) are constructed from near infrared (NIR)-emissive organic dyes with narrow absorption and emission bands and small Stokes shifts prone to aggregation-induced self-quenching. Here, we present the new asymmetric cyanine Itrybe with broad, almost environment-insensitive absorption and emission bands in the diagnostic window, offering a unique flexibility of the choice of excitation and detection wavelengths compared to common NIR dyes. This strongly emissive dye was spectroscopically studied in different solvents and encapsulated into differently sized (15, 25, 100 nm) amino-modified polystyrene NPs (PSNPs) via a one-step staining procedure. As proof-of-concept for its potential for pre-/clinical imaging applications, Itrybe-loaded NPs were surface-functionalized with polyethylene glycol (PEG) and the tumor-targeting antibody Herceptin and their binding specificity to the tumor-specific biomarker HER2 was systematically assessed. Itrybe-loaded NPs display strong fluorescence signals in vitro and in vivo and Herceptin-conjugated NPs bind specifically to HER2 as demonstrated in immunoassays as well as on tumor cells and sections from mouse tumor xenografts in vitro. This demonstrates that our design strategy exploiting broad band-absorbing and -emitting dyes yields versatile and bright NIR probes with a high potential for e.g. the sensitive detection and characterization of tumor development and progression. Oxford Elsevier 2013 Biomaterials 34 1 160 170 10.1016/j.biomaterials.2012.09.028 2016-02-19 OPUS4-27071 Posterpräsentation Behnke, Thomas Simple Approaches to Fluorescence Lifetime Standards Using Dye-Quencher Pairs 2012 BMT 2012 - Tagung der Biomedizinischen Technik BMT 2012 - Tagung der Biomedizinischen Technik Jena, Germany 2012-09-16 2012-09-19 2016-02-19 OPUS4-25481 Zeitschriftenartikel Behnke, Thomas; Würth, Christian; Laux, Eva-Maria; Hoffmann, Katrin; Resch-Genger, Ute Simple strategies towards bright polymer particles via one-step staining procedures In order to develop simple and versatile procedures for the preparation of red emissive particles, various one-step swelling procedures for the loading of fluorophores into nanometer- and micrometer-sized polystyrene particles were systematically assessed. Parameters studied for model dyes from common dye classes include the composition of the swelling medium, dye charge and polarity, dye concentration, and particle surface chemistry. The dye loading procedures were compared based upon the efficiency of dye incorporation, fluorescence intensity, and colloidal stability of the resulting particles as well as the absence of dye leaking as determined by absorption and fluorescence spectroscopy, flow cytometry, and measurements of zeta potentials. In addition, for the first time, the influence of the amount of incorporated dye on the absolute fluorescence quantum yield and brightness of the fluorescent particles was investigated for selected chromophores in differently sized particles using a custom-made calibrated integrating sphere setup. Our results demonstrate the general suitability of these one-step loading procedures for efficient particle staining with neutral, zwitterionic, and charged fluorophores like oxazines, coumarines, squaraines, xanthenes, and cyanines emitting in the visible and near infrared. Dye polarity was identified as a suitable tool to estimate the loading efficiency of fluorophores into these polymer particles. Kidlington Elsevier Ltd. 2012 Dyes and pigments 94 2 247 257 10.1016/j.dyepig.2012.01.021 2016-02-19 OPUS4-26051 Zeitschriftenartikel Laux, Eva-Maria; Behnke, Thomas; Hoffmann, Katrin; Resch-Genger, Ute Keeping particles brilliant - simple methods for the determination of the dye content of fluorophore-loaded polymeric particles One of the most active research areas in the life and material sciences is the design and synthesis of fluorescent nano- and micrometre sized particles for applications e.g. as labels, sensor systems, and platforms for fluorescence assays or barcoding materials. The reliable and reproducible fabrication of such particles as well as many applications require accurate, simple, and versatile procedures for the determination of the dye content per particle which affects e.g. the brightness of these materials and their surface charge and thus, colloidal stability. Here, four fast and inexpensive spectroscopic methods for the quantification of the fluorophore content of beads are presented and compared for nanometre- and micrometre sized polystyrene particles loaded or labeled with commercial fluorophores, differing in dye class, charge, and hydrophilicity. This included the determination of the amount of incorporated dye from absorption spectra of bead suspensions, via dissolving of the polymer matrix, via extraction of the polymer matrix, and from the supernatant of the swelling solution or reaction mixture. Method validation was performed with a sulfur-containing dye and elemental analysis. Based upon this method comparison and the accomplishable uncertainties, two reliable strategies for particle characterization and bead process control are identified that can be easily extended to other materials. Cambridge RSC Publ. Royal Society of Chemistry 2012 Analytical methods 4 6 1759 1768 10.1039/c2ay05822g 2016-02-19 OPUS4-30077 Zeitschriftenartikel Hoffmann, Katrin; Behnke, Thomas; Grabolle, Markus; Resch-Genger, Ute Nanoparticle-encapsulated vis- and NIR-emissive fluorophores with different fluorescence decay kinetics for lifetime multiplexing Bioanalytical, clinical, and security applications increasingly require simple, efficient, and versatile strategies to measure an ever increasing number of analytes or events in parallel in a broad variety of detection formats as well as in conjunction with chromatographic separation techniques or flow cytometry. An attractive alternative to common optical multiplexing and encoding methods utilizing spectral multiplexing/color encoding and intensity encoding is lifetime multiplexing, which relies on the discrimination between different fluorescent reporters based on their fluorescence decay kinetics. Here, we propose a platform of surface-functionalizable polymeric nanoparticles stained with fluorophores differing in their fluorescence lifetimes as a new multiplexing and encoding approach. Proof-of-concept measurements with different sets of lifetime-encoded polystyrene nanoparticles are presented, obtained via staining of preformed particles with visible (vis)- and near-infrared (NIR)-emissive organic dyes, which display very similar absorption and emission spectra to enable excitation and detection at the same wavelengths, yet sufficiently different fluorescence decay kinetics in suspension, thereby minimizing instrumentation costs. Data analysis was performed with a linear combination approach in the lifetime domain. Our results and first cell experiments with these reporter sets underline the suitability of our multiplexing strategy for the discrimination between and the quantification of different labels. This simple and versatile concept can be extended to all types of fluorophores, thereby expanding the accessible time scale, and can be used, e.g., for the design of labels and targeted probes for fluorescence assays and molecular imaging, cellular imaging studies, and barcoding applications, also in conjunction with spectral and intensity encoding. Berlin Springer 2014 Analytical and bioanalytical chemistry 406 14 3315 3322 10.1007/s00216-013-7597-3 2016-02-20 OPUS4-37125 Zeitschriftenartikel Schneider, Ralf; Weigert, Florian; Lesnyak, V.; Leubner, S.; Lorenz, T.; Behnke, Thomas; Dubavik, A.; Joswig, J.-O.; Resch-Genger, Ute; Gaponik, N.; Eychmüller, A. Resch-Genger, Ute; Schneider, Ralf pH and concentration dependence of the optical properties of thiol-capped CdTe nanocrystals in water and D2O The optical properties of semiconductor nanocrystals (SC NCs) are largely controlled by their size and surface chemistry, i.e., the chemical composition and thickness of inorganic passivation shells and the chemical nature and number of surface ligands as well as the strength of their bonds to surface atoms. The latter is particularly important for CdTe NCs, which - together with alloyed CdₓHg₁₋ₓTe - are the only SC NCs that can be prepared in water in high quality without the need for an additional inorganic passivation shell. Aiming at a better understanding of the role of stabilizing ligands for the control of the application-relevant fluorescence features of SC NCs, we assessed the influence of two of the most commonly used monodentate thiol ligands, thioglycolic acid (TGA) and mercaptopropionic acid (MPA), on the colloidal stability, photoluminescence (PL) quantum yield (QY), and PL decay behavior of a set of CdTe NC colloids. As an indirect measure for the strength of the coordinative bond of the ligands to SC NC surface atoms, the influence of the pH (pD) and the concentration on the PL properties of these colloids was examined in water and D₂O and compared to the results from previous dilution studies with a set of thiol-capped Cd₁₋ₓHgₓTe SC NCs in D₂O. As a prerequisite for these studies, the number of surface ligands was determined photometrically at different steps of purification after SC NC synthesis with Ellman's test. Our results demonstrate ligand control of the pH-dependent PL of these SC NCs, with MPA-stabilized CdTe NCs being less prone to luminescence quenching than TGA-capped ones. For both types of CdTe colloids, ligand desorption is more pronounced in H₂O compared to D₂O, underlining also the role of hydrogen bonding and solvent molecules. Cambridge RSC RSC 2016 Physical Chemistry Chemical Physics 18 28 19083 19092 urn:nbn:de:kobv:b43-371253 10.1039/c6cp03123d http://creativecommons.org/licenses/by-nc/3.0/de/deed.de 2016-08-12 OPUS4-31613 Vortrag Behnke, Thomas Design and Characterization of Streptavidin Modified Fluorescent Nanoparticles 2014 Molekulare Bildgebung 2014 Molekulare Bildgebung 2014 Jena, Germany 2014-10-08 2014-10-10 2016-02-20 OPUS4-21595 Zeitschriftenartikel Resch-Genger, Ute; Hoffmann, Katrin; Würth, Christian; Behnke, Thomas; Hoffmann, Angelika; Pfeifer, Dietmar; Engel, A. The toolbox of fluorescence standards: Flexible calibration tools for the standardization of fluorescence-based measurements To improve the reliability of fluorescence data in the life and material sciences and to enable accreditation of fluorescence techniques, standardization concepts are required that guarantee and improve the comparability of fluorescence measurements. At the core of such concepts are simple and evaluated fluorescence standards for the consideration of instrument-specific spectral and intensity distortions of measured signals and for instrument performance validation (IPV). Similarly in need are fluorescence intensity standards for the quantification from measured intensities and for signal referencing, thereby accounting for excitation light-induced intensity fluctuations. These standards should be preferably certified, especially for use in regulated areas like medical diagnostics. This encouraged us to develop liquid and solid standards for different fluorescence parameters and techniques for use under routine measurement conditions in different formates. Special emphasis was dedicated to the determination and control of the spectral responsivity of detection systems, wavelength accuracy, homogeneity of illumination, and intensity referencing for e.g. spectrofluorometers, fluorescence sensors and confocal laser scanning fluorescence microscopes. Here, we will present design concepts and examples for mono- and multifunctional fluorescence standards that provide traceability to radiometric units and present a first step towards a toolbox of standards. Redondo Beach, Calif. [u.a.] Soc. Society of Photo-Optical Instrumentation Engineers 2010 Proceedings / SPIE : P 7666 76661J 1 12 10.1117/12.853133 2016-02-19 OPUS4-21550 Posterpräsentation Behnke, Thomas Preparation, Characterization, and Bioanalytical Applications of Fluorescent Nano- and Micrometer-sized Particles 2010 Tag der Chemie Tag der Chemie Berlin, Germany 2010-06-16 2016-02-19 OPUS4-19418 Posterpräsentation Behnke, Thomas Encapsulated NIR Chromophores as Intensity Standards for Biosensor Systems 2009 ANAKON 2009 ANAKON 2009 Berlin, Germany 2009-03-17 2009-03-20 2016-02-19 OPUS4-19420 Posterpräsentation Behnke, Thomas NIR-Fluorescent Particles as Internal Standards in Biosensors 2009 Doktorandentagung Doktorandentagung Attendorn, Germany 2009-02-15 2009-02-17 2016-02-19 OPUS4-20423 Posterpräsentation Behnke, Thomas Encapsulation of NIR dyes: Comparison of different dye classes, spectroscopic evaluation, and stability studies 2009 11th International Conference on Methods and Applications of Fluorescence: Spectroscopy, Imaging and Probes 11th International Conference on Methods and Applications of Fluorescence: Spectroscopy, Imaging and Probes Budapest, Hungary 2009-09-06 2009-09-09 2016-02-19 OPUS4-23635 Beitrag zu einem Tagungsband Resch-Genger, Ute; Behnke, Thomas; Brehm, Robert; Grabolle, Markus; Hennig, Andreas; Hoffmann, Angelika; Hoffmann, Katrin; Linck, Lena; Lochmann, Cornelia; Pauli, Jutta; Spieles, Monika; Würth, Christian Funktionelle Chromophor-Systeme, innovative Validierungskonzepte und rückführbare Standards für die fluoreszenzbasierte multiparametrische Bioanalytik Unter dem Motto „Innovation und Qualitätssicherung in der (Bio)Analytik" werden in der Arbeitsgruppe Fluoreszenzspektroskopie der BAM, Bundesanstalt für Materialforschung und -prüfung, funktionelle Chromophor-Systeme, einfache Signalverstärkungs- und Multiplexingstrategien sowie innovative Validierungs- und rückführbare Standardisierungskonzepte für verschiedene fluorometrische Messgrößen und Methoden entwickelt. Im Mittelpunkt stehen dabei molekulare Fluorophore, Nanokristalle mit größenabhängigen optischen Eigenschaften (sogenannte Quantenpunkte, QDs) und fluoreszierende Partikel variabler Größe sowie Sonden und Sensormoleküle für neutrale und ionische Analyte und für die Charakterisierung von funktionellen Gruppen. Dabei erfolgen auch methodische Entwicklungen für die Fluoreszenzspektroskopie, die Fluoreszenzmikroskopie, die Milcrofluorometrie, die Sensorik und die Mikroarraytechnologie. Ziele sind u. a. das Design und die Untersuchung von multiplexfähigen selektiven und sensitiven Sonden für die Biomarkeranalytik, die Entwicklung von Methoden zur Charakterisierung der signalrelevanten Eigenschaften dieser Chromophor-Systeme und zur Charakterisierung von funktionellen Gruppen an Oberflächen und ihre Validierung sowie die Entwicklung und Bereitstellung von formatadaptierbaren, flexibel ersetzbaren Standards für die fluoreszenzbasierte Multiparameteranalytik. Senftenberg Hochschule Lausitz 2011 5. Senftenberger Innovationsforum Multiparameteranalytik (Proceedings) 5. Senftenberger Innovationsforum Multiparameteranalytik Senftenberg, Deutschland 10.03.2011 10.03.2011 86 108 2016-02-19 OPUS4-23637 Zeitschriftenartikel Hoffmann, Katrin; Behnke, Thomas; Drescher, Daniela; Kneipp, Janina; Resch-Genger, Ute Lifetime-based discrimination between spectrally matching vis and NIR emitting particle labels and probes Increasing the information content from bioassays which requires robust and efficient strategies for the detection of multiple analytes or targets in a single measurement is an important field of research, especially in the context of meeting current security and health concerns. An attractive alternative to spectral multiplexing, which relies on fluorescent labels excitable at the same wavelength, yet sufficiently differing in their emission spectra or color presents lifetime multiplexing. For this purpose, we recently introduced a new strategy based on 'pattern-matching' in the lifetime domain, which was exemplary exploited for the discrimination between organic dyes and quantum dot labels revealing multi-exponential decay kinetics and allowed quantification of these labels. Meanwhile, we have succeeded in extending this lifetime multiplexing approach to nanometer-sized particle labels and probes absorbing and emitting in the visible (vis) and near-infrared (NIR) spectral region. Here, we present a first proof-of-principle of this approach for a pair of NIR-fluorescent particles. Each particle is loaded with a single organic dye chosen to display very similar absorption and emission spectra, yet different fluorescence decay kinetics. Examples for the lifetime-based distinction between pairs of these fluorescent nanoparticles in solution and in cells are presented. The results underline the potential of fluorescenc lifetime multiplexing in life science and bioanalysis. Bellingham, Wash. SPIE, The International Society for Optical Engineering 2011 Proceedings of SPIE - Progress in biomedical optics and imaging 7905 79051F-1 79051F-9 10.1117/12.881442 2016-02-19 OPUS4-22184 Zeitschriftenartikel Würth, Christian; Hoffmann, Katrin; Behnke, Thomas; Ohnesorge, Marius; Resch-Genger, Ute Polymer- and glass-based fluorescence standards for the near infrared (NIR) spectral region The widespread use and acceptance of fluorescence techniques especially in regulated areas like medical diagnostics is closely linked to standardization concepts that guarantee and improve the comparability and reliability of fluorescence measurements. At the core of such concepts are dependable fluorescence standards that are preferably certified. The ever rising interest in fluorescence measurements in the near-infrared (NIR) spectral region renders the availability of spectral and intensity standards for this wavelength region increasingly important. This encouraged us to develop approaches to solid NIR standards based upon dye-doped polymers and assess their applicationrelevant properties in comparison to metal ion-doped glasses. The overall goal is here to provide inexpensive, easily fabricated, and robust internal and external calibration tools for a broad variety of fluorescence instruments ranging e.g. from spectrofluorometers over fluorescence microscopes to miniaturized fluorescence sensors. New York, NY Plenum Publ. Corp. 2011 Journal of fluorescence 21 3 953 961 10.1007/s10895-010-0650-0 2016-02-19 OPUS4-33695 Vortrag Behnke, Thomas FLIM and Optical Imaging with NIR-Fluorescent Lifetime-Encoded Nanoparticles of well Known Surface Chemistry 2015 Anakon 2015 Anakon 2015 Graz, Austria 2015-03-23 2015-03-26 2016-02-20 OPUS4-32406 Zeitschriftenartikel Würth, Christian; Geißler, Daniel; Behnke, Thomas; Kaiser, Martin; Resch-Genger, Ute Critical review of the determination of photoluminescence quantum yields of luminescent reporters A crucial variable for methodical performance evaluation and comparison of luminescent reporters is the photoluminescence quantum yield (Φ pl). This quantity, defined as the number of emitted photons per number of absorbed photons, is the direct measure of the efficiency of the conversion of absorbed photons into emitted light for small organic dyes, fluorescent proteins, metal-ligand complexes, metal clusters, polymeric nanoparticles, and semiconductor and up-conversion nanocrystals. Φ pl determines the sensitivity for the detection of a specific analyte from the chromophore perspective, together with its molar-absorption coefficient at the excitation wavelength. In this review we discuss different optical and photothermal methods for measuring Φ pl of transparent and scattering systems for the most common classes of luminescent reporters, and critically evaluate their potential and limitations. In addition, reporter-specific effects and sources of uncertainty are addressed. The ultimate objective is to provide users of fluorescence techniques with validated tools for the determination of Φ pl, including a series of Φ pl standards for the ultraviolet, visible, and near-infrared regions, and to enable better judgment of the reliability of literature data. Berlin Springer 2015 Analytical and bioanalytical chemistry 407 1 59 78 10.1007/s00216-014-8130-z 2016-02-20 OPUS4-36618 Zeitschriftenartikel Quevedo, Pablo; Behnke, Thomas; Resch-Genger, Ute Streptavidin conjugation and quantification—a method evaluation for nanoparticles Aiming at the development of validated protocols for protein conjugation of nanomaterials and the determination of protein labeling densities, we systematically assessed the conjugation of the model protein streptavidin (SAv) to 100-, 500-, and 1000-nm-sized polystyrene and silica nanoparticles and dye-encoded polymer particles with two established conjugation chemistries, based upon achievable coupling efficiencies and labeling densities. Bioconjugation reactions compared included EDC/sulfo NHS ester chemistry for direct binding of the SAv to carboxyl groups at the particle surface and maleimide-thiol chemistry in conjunction with heterobifunctional PEG linkers and aminated nanoparticles (NPs). Quantification of the total and functional amounts of SAv on these nanomaterials and unreacted SAv in solution was performed with the BCA assay and the biotin-FITC (BF) titration, relying on different signal generation principles, which are thus prone to different interferences. Our results revealed a clear influence of the conjugation chemistry on the amount of NP crosslinking, yet under optimized reaction conditions, EDC/sulfo NHS ester chemistry and the attachment via heterobifunctional PEG linkers led to comparably efficient SAv coupling and good labeling densities. Particle size can obviously affect protein labeling densities and particularly protein functionality, especially for larger particles. For unstained nanoparticles, direct bioconjugation seems to be the most efficient strategy, whereas for dyeencoded nanoparticles, PEG linkers are to be favored for the prevention of dye-protein interactions which can affect protein functionality specifically in the case of direct SAv binding. Moreover, an influence of particle size on achievable protein labeling densities and protein functionality could be demonstrated. Springer 2016 Analytical and Bioanalytical Chemistry 408 15 4133 4149 10.1007/s00216-016-9510-3 2016-06-22 OPUS4-44038 Zeitschriftenartikel Kothavale, S.; Jadhav, A. G.; Scholz, Norman; Nirmalananthan-Budau, Nithiya; Behnke, Thomas; Resch-Genger, Ute; Sekar, N. Corrigendum to "Deep red emitting triphenylamine based coumarin-rhodamine hybrids with large stokes shift and viscosity sensing: Synthesis, photophysical properties and DFT studies of their spirocyclic and open forms" [Dyes Pigments 137 (2017) 329-341] We designed and synthesized triphenylamine based and coumarin fused rhodamine hybrid dyes and characterized using 1H, 13C NMR and HR-LCMS analysis. Both the newly synthesized hybrid dyes were found to show red shifted absorption as well as emissions and large Stokes shift (40e68 nm) as compared to the small Stokes shift (25e30 nm) of reported dyes Rhodamine B and 101. Photophysical properties of these dyes were studied in different solvents and according to the solvents acidity or basicity they preferred to remain in their spirocyclic or open form in different ratio. We studied the spirocyclic as well as open form derivatives of these dyes for their viscosity sensitivity in three different mixture of solvents i.e. polar-protic [EtOH-PEG 400], polar-aprotic [toluene-PEG 400] and non-polaraprotic [toluene-paraffin]. They are found to show very high viscosity sensitivity in polar-protic mixture of solvents [EtOH-PEG 400] and hence concluded that both polarity as well as viscosity factor worked together for the higher emission enhancement rather than only viscosity factor. As these dyes showed very high viscosity sensitivity in their spirocyclic as well as open form, they can be utilized as viscosity sensors in visible as well as deep red region. We also correlated our experimental finding theoretically by using Density Functional theory computations. Amsterdam Elsevier 2018 Dyes and pigments 149 929 10.1016/j.dyepig.2017.06.021 2018-02-08 OPUS4-44830 Zeitschriftenartikel Moser, Marko; Nirmalananthan-Budau, Nithiya; Behnke, Thomas; Geißler, Daniel; Resch-Genger, Ute Multimodal cleavable reporters versus conventional labels for optical quantification of accessible amino and carboxy groups on nano- and microparticles Many applications of nanometer- and micrometer-sized particles include their surface functionalization with linkers, sensor molecules, and analyte recognition moieties like (bio)ligands. This requires knowledge of the chemical nature and number of surface groups accessible for subsequent coupling reactions. Particularly attractive for the quantification of these groups are spectrophotometric and fluorometric assays, which can be read out with simple instrumentation. In this respect, we present here a novel Family of cleavable spectrophotometric and multimodal reporters for conjugatable amino and carboxyl surface groups on nano- and microparticles. This allows determination of particle-bound labels, unbound reporters in the supernatant, and reporters cleaved off from the particle surface, as well as the remaining thiol groups on particle, by spectrophotometry and inductively coupled optical emission spectrometry (32S ICP-OES). Comparison of the performance of these cleavable reporters with conductometry and conventional labels, utilizing changes in intensity or Color of absorption or emission, underlines the analytical potential of this versatile concept which elegantly circumvents Signal distortions by scattering and Encoding dyes and enables straightforward validation by method comparison. Washington, DC ACS Publ. 2018 Analytical chemistry 90 9 5887 5895 10.1021/acs.analchem.8b00666 2018-05-04 OPUS4-35302 Zeitschriftenartikel Markus, M. A.; Napp, J.; Behnke, Thomas; Mitkovski, M.; Monecke, S.; Dullin, C.; Kilfeather, S.; Dressel, R.; Resch-Genger, Ute; Alves, F. Tracking of inhaled near-infrared fluorescent nanoparticles in lungs of SKH-1 mice with allergic airway inflammation Molecular imaging of inflammatory lung diseases, such as asthma, has been limited to date. The recruitment of innate immune cells to the airways is central to the inflammation process. This study exploits these cells for imaging purposes within the lung, using inhaled polystyrene nanoparticles loaded with the near-infrared fluorescence dye Itrybe (Itrybe-NPs). By means of in vivo and ex vivo fluorescence reflectance imaging of an ovalbumin-based allergic airway inflammation (AAI) model in hairless SKH-1 mice, we show that subsequent to intranasal application of Itrybe-NPs, AAI lungs display fluorescence intensities significantly higher than those in lungs of control mice for at least 24 h. Ex vivo immunofluorescence analysis of lung tissue demonstrates the uptake of Itrybe-NPs predominantly by CD68+CD11c+ECF-L+MHCIIlow cells, identifying them as alveolar M2 macrophages in the peribronchial and alveolar areas. The in vivo results were validated by confocal microscopy, overlapping tile analysis, and flow cytometry, showing an amount of Itrybe-NP-containing macrophages in lungs of AAI mice significantly larger than that in controls. A small percentage of NP-containing cells were identified as dendritic cells. Flow cytometry of tracheobronchial lymph nodes showed that Itrybe-NPs were negligible in lung draining lymph nodes 24 h after inhalation. This imaging approach may advance preclinical monitoring of AAI in vivo over time and aid the investigation of the role that macrophages play during lung inflammation. Furthermore, it allows for tracking of inhaled nanoparticles and can hence be utilized for studies of the fate of potential new nanotherapeutics. Washington, DC, USA ACS Publ. American Chemical Society 2015 ACS nano 9 12 11642 11657 10.1021/acsnano.5b04026 2016-02-20 OPUS4-34468 Zeitschriftenartikel Moser, Marko; Behnke, Thomas; Hamers-Allin, Carolina; Klein-Hartwig, Karin; Falkenhagen, Jana; Resch-Genger, Ute Quantification of PEG-maleimide ligands and coupling efficiencies on nanoparticles with Ellman's reagent The surface modification of nanometer- and micrometer-sized particles and planar substrates with polyethylene glycol (PEG) ligands of varying length is a very common strategy to tune the hydrophilicity and biocompatibility of such materials, minimize unspecific interactions, improve biofunctionalization efficiencies, and enhance blood circulation times. Nevertheless, simple methods for the quantification of PEG ligands are comparatively rare. Here, we present a new concept for the quantification of PEG ligands for maleimide-functionalized PEG molecules and the determination of PEG coupling efficiencies, exploiting the quantitative reaction of maleimide with ʟ-cysteine, and the subsequent determination of the unreacted thiol with the photometric Ellman's test. This is shown for heterobifunctional PEG spacers of varying length and amino-functionalized polystyrene nanoparticles (PS NP) without and with differently charged encoding dyes. The reaction of ʟ-cysteine with the Ellman's reagent was monitored photometrically and with electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS) to derive the reaction mechanism and to obtain the stoichiometry factor for ʟ-cysteine quantification. Mass balances and quantification of ʟ-cysteine via its sulfur concentration using elemental analysis and inductively coupled plasma mass spectrometry (ICP-MS) confirmed the accuracy and reliability of this approach that can be extended to other surface groups and ligands. Washington, DC American Chemical Society 2015 Analytical chemistry 87 18 9376 9383 10.1021/acs.analchem.5b02173 2016-02-20 OPUS4-34723 Vortrag Behnke, Thomas Fluoreszente Polymer-Nanopartikel für die Bioanalytik 2015 Nano- und Mikropartikel für die biomedizinische Analytik Nano- und Mikropartikel für die biomedizinische Analytik Potsdam-Golm 2015-10-29 2016-02-20 OPUS4-43213 Zeitschriftenartikel Denißen, M.; Nirmalananthan-Budau, Nithiya; Behnke, Thomas; Hoffmann, Katrin; Resch-Genger, Ute; Müller, T. J. J. 3-Piperazinyl propenylidene indolone merocyanines: consecutive three-component synthesis and electronic properties of solid-state luminophores with AIE properties A series of twelve 3-piperazinyl propenylidene indolone merocyanines was synthesized in a one-pot fashion using a consecutive three-component insertion-coupling-Michael addition sequence. Physicalorganic treatment of the absorption data of a consanguineous series of this library allows semiquantitative Linear Free Energy Relationships (LFERs) to be established and confirmation of the positive Absorption solvatochromicity. All Boc-substituted piperazinyl merocyanines display aggregation induced Emission (AIE), which was corroborated for two solvent systems. In particular, crystallization-induced Emission enhancement (CIEE) induced by ultrasonication could be shown for a model chromophore by confocal laser scanning microscopy (CLSM). 2017 MATERIALS CHEMISTRY FRONTIERS 1 10 2013 2026 10.1039/c7qm00198c 2017-12-01 OPUS4-40004 Vortrag Behnke, Thomas Spectroscopic quantification of surface groups on micro- and nanoparticles Nanometer- and micrometer-sized particles are of increasing importance for a wide range of applications in the material and life sciences. This includes carriers for, e.g., drugs as well as dye molecules for use as multichromophoric reporters for signal enhancement in optical assays, platforms for DNA sequencing, and the fabrication of nanosensors and targeted probes for bioimaging studies. Application-relevant properties of such particles include their size (and size distribution), shape, colloidal stability, biocompatibility, and ease of subsequent functionalization, e.g., with linkers, sensor molecules, and targeting ligands. The latter requires knowledge of the number of groups effectively accessible for subsequent coupling reactions and hence, selective and sensitive methods of analysis, which can be ideally employed for the characterization of a broad variety of particle systems independent of their optical properties, i.e., scattering or the presence of encoding dyes. For product control, robust, reliable and fast methods performable with inexpensive equipment are prefered. In this respect, we studied a variety of conventional labels for optical readout, utilizing changes in intensity and/or color of absorption and/or emission. In addition, we developed a platform of cleavable and multimodal labels for optical assays which consist of a cleavable linker and an optically active reporter moiety. In contrast to conventional reporters measured directly at the particle surface, which are prone to signal distortions by scattering and encoding dyes, these cleavable labels can be detected colorimetrically or fluorometrically both bound at the particle surface and after quantitative cleavage of the linker in the transparent supernatant. Moreover, they enable straightforward validation by method comparison with elemental analysis, ICP-OES or ICP-MS. Here, we present representative examples of newly synthesized cleavable labels and their application for the quantification of amino, thiol and carboxy surface groups on different nanomaterials and compare these results with measurements using conventional optical labels. 2017 253rd ACS National Meeting San Francisco, CA, USA 02.04.2017 06.04.2017 2017-04-28 OPUS4-39211 Zeitschriftenartikel Gers-Panther, C.F.; Fischer, H.; Nordmann, J.; Seiler, T.; Behnke, Thomas; Würth, Christian; Frank, W.; Resch-Genger, Ute; Müller, T.J.J. Four- and Five-Component Syntheses and Photophysical Properties of Emission Solvatochromic 3‑Aminovinylquinoxalines 3-Aminovinylquinoxalines are readily accessible from (hetero)aryl glyoxylic acids or heterocyclic π-nucleophiles by consecutive four- and fivecomponent syntheses in the sense of an activation-alkynylation-cyclocondensation-addition sequence or glyoxylation-alkynylation-cyclocondensation-addition sequence in good yields. The title compounds are highly fluorescent with pronounced emission solvatochromicity and protochromic fluorescence quenching. Time-resolved fluorescence spectroscopy furnishes radiative and nonradiative fluorescence decay rates in various solvent polarities. The electronic structure is corroborated by DFT and TD-DFT calculations rationalizing the observed spectroscopic effects. ACS Publications 2016 Journal of organic chemistry 82 567 578 10.1021/acs.joc.6b02581 2017-02-22 OPUS4-42584 Zeitschriftenartikel Scholz, Norman; Jadhav, A.; Shreykar, M.; Behnke, Thomas; Nirmalananthan-Budau, Nithiya; Resch-Genger, Ute; Sekar, N. Coumarin-rhodamine hybrids-novel probes for the optical measurements of viscosity and polarity A comprehensive systematic study of Absorption and fluorescence properties in solvents of varying viscosity and polarity of three novel and red-emitting coumarin-rhodamine hybrid derivatives with differences in the rigidity of their substituents is presented. This includes ethanol-polyethylene glycol, toluene-polyethylene glycol, and tolueneparaffin mixtures. Moreover, protonation-induced effects on the spectroscopic properties are studied. A viscosity-induced emission enhancement was observed for all coumarin-rhodamine hybrid derivatives. MCR2 bearing a julolidine donor showed the expected low sensitivity to viscosity whereas MCR3 with its freely rotatable diphenylamino Substituent revealed a particularly pronounced sensitivity to this parameter. Moreover, MCR2 shows an enhancement in Emission in the open, i.e., protonated form in conjunction with a largely Stokes shift fluorescence in the deep red spectral region. This enables the application of these dyes as viscosity sensors and as far red emitting pH-sensitive probes. CrossMark Springer Science+Business Media 2017 Journal of Fluorescence 27 6 1949 1956 10.1007/s10895-017-2165-4 2017-10-20 OPUS4-41783 Zeitschriftenartikel Pauli, Jutta; Hoffmann, Katrin; Würth, Christian; Behnke, Thomas; Resch-Genger, Ute Standardization of fluorescence measurements in the UV/vis/NIR/IR Photoluminescence techniques are amongst the most widely used Tools in the life sciences, with new and exciting applications in medical diagnostics and molecular Imaging continuously emerging. Advantages include their comparative ease of use, unique sensitivity, non-invasive character, and potential for Multiplexing, remote sensing, and miniaturization. General drawbacks are, however, signals, that contain unwanted wavelength- and polarization contributions from Instrument-dependent effects, which are also time-dependent due to aging of Instrument-components, and difficulties to measure absolute flourescence entensities. Moreover, scattering Systems require Special measurement geometries and the interest in new optical Reporters with Emission > 1000 nm strategies for reliable measurements in the second diagnostic for the comparison of material Performance and the rational designg of new flourophores with improved properties. Here, we present strategies to versatile method-adaptable liquid and solid flourescence Standards for different flourescence paramters including traceable Instrument calibration procedures and the design of integrating spere setups for the absolute measurements of emission spectra and Quantum yields in the wavelength Region of 350 to 1600 nm. Examples are multi-Emitter glasses, spectral flourescence Standards, and quantum yield Standards for the UV/vis/NIR. Proceedings of SPIE 2017 Standardization of Fluorescence Measurements in the UV/vis/NIR/IR 10049 978-1-5106-0539-8 Conference on Molecular-Guided Surgery - Molecules, Devices, and Applications III San Francisco, CA, USA 28.01.2017 30.01.2017 1 10.1117/12.2255728 2017-11-06 OPUS4-57368 Zeitschriftenartikel Würth, Christian; Behnke, Thomas; Gienger, J.; Resch-Genger, Ute Efficiency scale for scatteringluminescent particles linkedto fundamental and measurablespectroscopic properties Comparing the performance of molecular and nanoscale luminophores and luminescent microand nanoparticles and estimating achievable signal amplitudes and limits of detection requires a standardizable intensity scale. This initiated the development of the relative MESF (number of molecules of equivalent soluble fluorochromes) and ERF (equivalent reference fluorophores) scales for flow cytometry and fluorescence microscopy. Both intensity scales rely on fluorescence intensity values assigned to fluorescent calibration beads by an intensity comparison to spectrally closely matching fluorophore solutions of known concentration using a spectrofluorometer. Alternatively, the luminophore or bead brightness (B) can be determined that equals the product of the absorption cross section (σa) at the excitation wavelength (σa(λex)) and the photoluminescence quantum yield (Φpl). Thereby, an absolute scale based on fundamental and measurable spectroscopic properties can be realized which is independent of particle size, material, and luminophore staining or labeling density and considers the sensitivity of the optical properties of luminophores to their environment. Aiming for establishing such a brightness scale for light-scattering dispersions of luminescent particles with sizes exceeding a few ten nanometers, we demonstrate how the brightness of quasi-monodisperse 25 nm, 100 nm, and 1 μm sized polystyrene particles (PSP), loaded with two different dyes in varying concentrations, can be obtained with a single custom-designed integrating sphere setup that enables the absolute determination of Φpl and transmittance and diffuse reflectance measurements. The resulting Φpl, σa(λex), imaginary parts of the refractive index, and calculated B values of these samples are given in dependence of the number of incorporated dye molecule per particle. Finally, a unitless luminescence efficiency (LE) is defined allowing for the direct comparison of luminescence efficiencies of particles with different sizes. Nature 2023 Scientific Reports 13 1 14 urn:nbn:de:kobv:b43-573680 10.1038/s41598-023-32933-6 https://creativecommons.org/licenses/by/4.0/deed.de 2023-04-24