TY - JOUR A1 - Eisold, Ursula A1 - Sellrie, Frank A1 - Schenk, Jörg A. A1 - Lenz, Christine A1 - Stöcklein, Walter F. M. A1 - Kumke, Michael Uwe T1 - Bright or dark immune complexes of anti-TAMRA antibodies for adapted fluorescence-based bioanalysis JF - Analytical & bioanalytical chemistry N2 - Fluorescence labels, for example fluorescein or rhodamin derivatives, are widely used in bioanalysis applications including lateral-flow assays, PCR, and fluorescence microscopy. Depending on the layout of the particular application, fluorescence quenching or enhancement may be desired as the detection principle. Especially for multiplexed applications or high-brightness requirements, a tunable fluorescence probe can be beneficial. The alterations in the photophysics of rhodamine derivatives upon binding to two different anti-TAMRA antibodies were investigated by absorption and fluorescence-spectroscopy techniques, especially determining the fluorescence decay time and steady-state and time-resolved fluorescence anisotropy. Two monoclonal anti-TAMRA antibodies were generated by the hybridoma technique. Although surface-plasmon-resonance measurements clearly proved the high affinity of both antibodies towards 5-TAMRA, the observed effects on the fluorescence of rhodamine derivatives were very different. Depending on the anti-TAMRA antibody either a strong fluorescence quenching (G71-DC7) or a distinct fluorescence enhancement (G71-BE11) upon formation of the immune complex was observed. Additional rhodamine derivatives were used to gain further information on the binding interaction. The data reveal that such haptens as 5-TAMRA could generate different paratopes with equal binding affinities but different binding interactions, which provide the opportunity to adapt bioanalysis methods including immunoassays for optimized detection principles for the same hapten depending on the specific requirements. KW - mAb KW - Fluorescence KW - Anisotropy KW - Exciplex KW - Energy-transfer probe Y1 - 2015 U6 - https://doi.org/10.1007/s00216-015-8538-0 SN - 1618-2642 SN - 1618-2650 VL - 407 IS - 12 SP - 3313 EP - 3323 PB - Springer CY - Heidelberg ER - TY - JOUR A1 - Haubitz, Toni A1 - Fudickar, Werner A1 - Linker, Torsten A1 - Kumke, Michael Uwe T1 - pH-sensitive fluorescence switching of pyridylanthracenes BT - the effect of the isomeric pattern JF - The journal of physical chemistry : A, Molecules, spectroscopy, kinetics, environment & general theory N2 - 9,10-substituted anthracenes are known for their useful optical properties like fluorescence, which makes them frequently used probes in sensing applications. In this article, we investigate the fundamental photophysical properties of three pyridyl-substituted variants. The nitrogen atoms in the pyridinium six-membered rings are located in the ortho-, meta-, and para-positions in relation to the anthracene core. Absorption, fluorescence, and transient absorption measurements were carried out and were complemented by theoretical calculations. We monitored the photophysics of the anthracene derivatives in chloroform and water investigating the protonated as well as their nonprotonated forms. We found that the optical properties of the nonprotonated forms are strongly determined by the anthracene chromophore, with only small differences to other 9,10-substituted anthracenes, for example diphenyl anthracene. In contrast, protonation leads to a strong decrease in fluorescence intensity and lifetime. Transient absorption measurements and theoretical calculations revealed the formation of a charge-transfer state in the protonated chromophores, where electron density is shifted from the anthracene moiety toward the protonated pyridyl substituents. While the para- and ortho-derivatives' charge transfer is still moderately fluorescent, the meta-derivative is affected much stronger and shows nearly no fluorescence. This nitrogen-atom-position-dependent sensitivity to hydronium activity makes a combination of these fluorophores very attractive for pH-sensing applications covering a broadened pH range. KW - Absorption KW - Aromatic compounds KW - Fluorescence KW - Hydrocarbons KW - Reaction mechanisms Y1 - 2020 U6 - https://doi.org/10.1021/acs.jpca.0c09911 SN - 1089-5639 SN - 1520-5215 VL - 124 IS - 52 SP - 11017 EP - 11024 PB - American Chemical Society CY - Washington ER - TY - JOUR A1 - Kienzler, Andrea Altevogt Nee A1 - Flehr, Roman A1 - Gehne, Sören A1 - Kumke, Michael Uwe A1 - Bannwarth, Willi T1 - Verification and biophysical characterization of a New Three-Color Forster Resonance-Energy-Transfer (FRET) System in DNA JF - Helvetica chimica acta N2 - We report on a new three-color FRET system consisting of three fluorescent dyes, i.e., of a carbostyril (=quinolin-2(1H)-one)-derived donor D, a (bathophenanthroline)ruthenium complex as a relay chromophore A1, and a Cy dye as A2 (FRET=Forster resonance-energy-transfer) (cf. Fig. 1). With their widely matching spectroscopic properties (cf. Fig. 2), the combination of these dyes yielded excellent FRET efficiencies. Furthermore, fluorescence lifetime measurements revealed that the long fluorescence lifetime of the Ru complex was transferred to the Cy dye offering the possibility to measure the whole system in a time-resolved mode. The FRET system was established on double-stranded DNA (cf. Fig. 3) but it should also be generally applicable to other biomolecules. KW - Forster resonance energy transfer (FRET) system KW - DNA KW - Fluorescence KW - Ruthenium complexes Y1 - 2012 U6 - https://doi.org/10.1002/hlca.201100460 SN - 0018-019X VL - 95 IS - 4 SP - 543 EP - 555 PB - Wiley-VCH CY - Weinheim ER - TY - THES A1 - Kumke, Michael Uwe T1 - Huminstoffe und organische Modellliganden und ihre Wechselwirkung mit Metallionen und polyzyklischen aromatischen Kohlenwasserstoffen T1 - Humic substances and organic model ligands – Interactions with metal ions and polycyclic aromatic hydrocarbons N2 - Immobilisierung bzw. Mobilisierung und Transport von Schadstoffen in der Umwelt, besonders in den Kompartimenten Boden und Wasser, sind von fundamentaler Bedeutung für unser (Über)Leben auf der Erde. Einer der Hauptreaktionspartner für organische und anorganische Schadstoffe (Xenobiotika) in der Umwelt sind Huminstoffe (HS). HS sind Abbauprodukte pflanzlichen und tierischen Gewebes, die durch eine Kombination von chemischen und biologischen Ab- und Umbauprozessen entstehen. Bedingt durch ihre Genese stellen HS außerordentlich heterogene Stoffsysteme dar, die eine Palette von verschiedenartigen Wechselwirkungen mit Schadstoffen zeigen. Die Untersuchung der fundamentalen Wechselwirkungsmechanismen stellt ebenso wie deren quantitative Beschreibung höchste Anforderungen an die Untersuchungsmethoden. Zur qualitativen und quantitativen Charakterisierung der Wechselwirkungen zwischen HS und Xenobiotika werden demnach analytische Methoden benötigt, die bei der Untersuchung von extrem heterogenen Systemen aussagekräftige Daten zu liefern vermögen. Besonders spektroskopische Verfahren, wie z.B. lumineszenz-basierte Verfahren, besitzen neben der hervorragenden Selektivität und Sensitivität, auch eine Multidimensionalität (bei der Lumineszenz sind es die Beobachtungsgrößen Intensität IF, Anregungswellenlänge lex, Emissionswellenlänge lem und Fluoreszenzabklingzeit tF), die es gestattet, auch heterogene Systeme wie HS direkt zu untersuchen. Zur Charakterisierung können sowohl die intrinsischen Fluoreszenzeigenschaften der HS als auch die von speziell eingeführten Lumineszenzsonden verwendet werden. In beiden Fällen werden die zu Grunde liegenden fundamentalen Konzepte der Wechselwirkungen von HS mit Xenobiotika untersucht und charakterisiert. Für die intrinsische Fluoreszenz der HS konnte gezeigt werden, dass neben molekularen Strukturen besonders die Verknüpfung der Fluorophore im Gesamt-HS-Molekül von Bedeutung ist. Konformative Freiheit und die Nachbarschaft zu als Energieakzeptor fungierenden HS-eigenen Gruppen sind wichtige Komponenten für die Charakteristik der HS-Fluoreszenz. Die Löschung der intrinsischen Fluoreszenz durch Metallkomplexierung ist demnach auch das Resultat der veränderten konformativen Freiheit der HS durch die gebundenen Metallionen. Es zeigte sich, dass abhängig vom Metallion sowohl Löschung als auch Verstärkung der intrinsischen HS-Fluoreszenz beobachtet werden kann. Als extrinsische Lumineszenzsonden mit wohl-charakterisierten photophysikalischen Eigenschaften wurden polyzyklische aromatische Kohlenwasserstoffe und Lanthanoid-Ionen eingesetzt. Durch Untersuchungen bei sehr niedrigen Temperaturen (10 K) konnte erstmals die Mikroumgebung von an HS gebundenen hydrophoben Xenobiotika untersucht werden. Im Vergleich mit Raumtemperaturexperimenten konnte gezeigt werden, dass hydrophobe Xenobiotika an HS-gebunden in einer Mikroumgebung, die in ihrer Polarität analog zu kurzkettigen Alkoholen ist, vorliegen. Für den Fall der Metallkomplexierung wurden Energietransferprozesse zwischen HS und Lanthanoidionen bzw. zwischen verschiedenen, gebundenen Lanthanoidionen untersucht. Basierend auf diesen Messungen können Aussagen über die beteiligten elektronischen Zustände der HS einerseits und Entfernungen von Metallbindungsstellen in HS selbst angeben werden. Es ist dabei zu beachten, dass die Experimente in Lösung bei realen Konzentrationen durchgeführt wurden. Aus Messung der Energietransferraten können direkte Aussagen über Konformationsänderungen bzw. Aggregationsprozesse von HS abgeleitet werden. N2 - Transport and fate of xenobiotics in the environment, especially in water and soil, are of utmost importance for life on earth. A major reaction partner for xenobiotics in the environment are humic substances (HS). HS are degradation products of plant and animal tissue, which are formed in a combination of subsequent chemical and/or biochemical processes. Because of the complex history of their origin HS are extremely heterogeneous mixtures of different compounds. Consequently, they posses a great variety of interaction capabilities with various xenobiotics. The investigations of the fundamental interaction mechanisms between HS and xenobiotics make high demands on the analytical techniques used. Especially spectroscopic techniques are promising for the investigation of interaction mechanisms in complex systems. Luminescence spectroscopy has the great advantage of outstanding sensitivity and of multidimensionality, which in principle allows the investigation of HS under environmental relevant conditions. For the characterisation of interaction processes of HS with xenobiotics the intrinsic fluorescence of HS as well as the luminescence of extrinsic probes can be used. The intrinsic HS fluorescence is determined by the molecular structure as well as the connection of the basic fluorophores. Conformational freedom as well as the presence of energy accepting groups in the neighbourhood of the fluorophores are highly important for the overall intrinsic HS fluorescence. The presence of metal ions can either quench or enhance the intrinsic HS fluorescence, which depends on the metal ion as well as on the origin of the HS investigated. While in most cases Al3+ ions enhance the intrinsic HS fluorescence, Ln3+ ions induce a fluorescence quenching. Polycyclic aromatic hydrocarbons were used as extrinsic fluorescence probes in order to characterize the interaction of HS and hydrophobic organic xenobiotics. In investigations at ultra-low temperatures (10 K) it could be shown that pyrene is bound in a HS microenvironment with an polarity which resembles that of small alcohols (e.g., butanol). In case of metal complexation, the lanthanide ions Eu3+ and Tb3+ were used as luminescence probes. Due to the outstanding luminescence properties of those ions, information about metal binding sites in HS were obtained. Based on the measurements of intramolecular and intermolecular energy transfer processes average distances of metal binding sites were deduced. KW - Fluoreszenz KW - Huminstoffe KW - Lanthanoide KW - Huminstoffe KW - Fluoreszenz KW - Lanthanoide KW - Resonanzenergietransfer KW - Lumineszenzsonden KW - Fluorescence KW - humic substances KW - resonance energy transfer KW - luminescence probes KW - lanthanides Y1 - 2005 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus-6066 ER - TY - JOUR A1 - Techen, Anne A1 - Czapla, Sylvia A1 - Möllnitz, Kristian A1 - Budach, Dennis B. A1 - Wessig, Pablo A1 - Kumke, Michael Uwe T1 - Synthesis and spectroscopic characterization of fluorophore-labeled oligospiroketal rods JF - Helvetica chimica acta N2 - Fluorescence probes consisting of well-established fluorophores in combination with rigid molecular rods based on spirane-type structures were investigated with respect to their fluorescence properties under different solvent conditions. The attachment of the dyes was accomplished by 1,3-dipolar cycloaddition between alkynes and azides (click' reaction) and is a prime example for a novel class of sensor constructs. Especially, the attachment of two (different) fluorophores on opposite sides of the molecular rods paves the way to new sensor systems with less bulky (compared to the conventional DNA- or protein-based concepts), nevertheless rigid spacer constructs, e.g., for FRET-based sensing applications. A detailed photophysical characterization was performed in MeOH (and in basic H2O/MeOH mixtures) for i) rod constructs containing carboxyfluorescein, ii) rod constructs containing carboxyrhodamine, iii) rod constructs containing both carboxyfluorescein and carboxyrhodamine, and iv) rod constructs containing both pyrene and perylene parts. For each dye (pair), two rod lengths with different numbers of spirane units were synthesized and investigated. The rod constructs were characterized in ensemble as well as single-molecule fluorescence experiments with respect to i) specific roddye and ii) dyedye interactions. In addition to MeOH and MeOH/NaOH, the rod constructs were also investigated in micellar systems, which were chosen as a simplified model for membranes. KW - Molecular rods KW - Forster resonance energy transfer (FRET) KW - Carboxyfluorescein KW - Carboxyrhodamine KW - Pyrene KW - Perylene KW - Fluorescence Y1 - 2013 U6 - https://doi.org/10.1002/hlca.201200616 SN - 0018-019X SN - 1522-2675 VL - 96 IS - 11 SP - 2046 EP - 2067 PB - Wiley-VCH CY - Weinheim ER - TY - JOUR A1 - Wessig, Pablo A1 - Behrends, Nicole A1 - Kumke, Michael Uwe A1 - Eisold, Ursula T1 - FRET Pairs with Fixed Relative Orientation of Chromophores JF - European journal of organic chemistry N2 - Synthetic routes to different oligospirothioketal (OSTK) Forster resonance energy transfer (FRET) constructs are described and the photophysics of these constructs were explored in different solvents. The FRET efficiencies were determined from the experimental data and compared with theoretical values. The influence of the outstanding rigidity of the novel OSTK compounds on the FRET is discussed. KW - Fluorescence KW - Energy transfer KW - FRET KW - Chromophores KW - Spiro compounds Y1 - 2016 U6 - https://doi.org/10.1002/ejoc.201600489 SN - 1434-193X SN - 1099-0690 VL - 145 SP - 4476 EP - 4486 PB - Wiley-VCH CY - Weinheim ER -