TY - CONF A1 - Linck, Lena A1 - Resch-Genger, Ute A1 - Reiß, Edda A1 - Bier, Frank A1 - Sobek, Jens T1 - Optimization of fluorescence signals on rolling amplification-enhanced microarrays T2 - Dechema Tagung functional genomics T2 - Dechema Tagung functional genomics CY - Frankfurt am Main, Germany DA - 2011-02-03 PY - 2011 AN - OPUS4-23536 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Linck, Lena A1 - Resch-Genger, Ute T1 - Optimized Fluorescence Detection for RCA-Assays on Microarrays T2 - Kolloquium T2 - Kolloquium CY - Potsdam, Germany DA - 2010-08-25 PY - 2010 AN - OPUS4-22081 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Linck, Lena A1 - Kapusta, P. A1 - Resch-Genger, Ute T1 - Spectroscopic and photophysical properties of dUTP and internally DNA bound fluorophores for optimized signal detection in biological formats JF - Photochemistry and photobiology N2 - Efficient signal generation in DNA-based assays requires understanding of the influence of fluorophore’s interactions on the spectroscopic properties. The resulting changes in fluorescence intensity, quantum yield, emission anisotropy, and fluorescence lifetime provide straightforward tools for the study of molecular dynamics and interaction between labels and nucleic acids. Searching for bright fluorescent reporters for rolling circle amplification (RCA) as efficient signal enhancement strategy for biological formats, we investigated the spectroscopic properties of seven dyes: cyanines, rhodamines, and BODIPYs. They spectrally resemble Cy3, the most frequently used fluorophore in biodetection formats, and are measured in six samples (free dye, dye-dUTP, internally labeled ssDNA and dsDNA—single- and triple-labeled) using steady-state and time-resolved fluorometry. Special emphasis was dedicated to characterizing the nature of the interaction of these fluorophores differing in dye class, charge, and rigidity. Our results suggest dye charge and structure as main factors governing the dye’s interactions, with DY-555 and Cy3B presenting the best candidates for our envisaged signal amplification strategy. This label comparison underlines the importance of a proper understanding of structure-property relations and dye–biomolecule interactions for reporter choice and presents a road map towards the design and interpretation of experiments using these labels on DNA of known sequence. KW - DNA KW - Fluorescent dye KW - Fluorescence quantum yield KW - Cyanine KW - Rhodamine KW - Fluorescence lifetime KW - Guanosine quenching PY - 2012 DO - https://doi.org/10.1111/j.1751-1097.2012.01119.x SN - 0031-8655 SN - 1751-1097 VL - 88 IS - 4 SP - 867 EP - 875 PB - American Society for Photobiology CY - Augusta, Ga. AN - OPUS4-27080 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Linck, Lena A1 - Reiß, E. A1 - Bier, F. A1 - Resch-Genger, Ute T1 - Direct labeling rolling circle amplification as a straightforward signal amplification technique for biodetection formats JF - Analytical methods N2 - Biodetection formats, such as DNA and antibody microarrays, are valuable tools in the life sciences, but for some applications, the detection limits are insufficient. A straightforward strategy to obtain signal amplification is the rolling circle amplification (RCA), an easy, isothermal, and enzymatic nucleic acid synthesis that has already been employed successfully to increase the signal yield for several single-analyte and multiplexing assays in conjunction with hybridization probes. Here, we systematically investigated the parameters responsible for the RCA driven signal amplification with fluorescent labels, such as the type of fluorophore chosen, labeling strategy, composition of reaction solution, and number of handling steps. In labeling strategies, post-synthetic labeling via a Cy3-hybridization probe was compared to the direct incorporation of fluorescent Cy3–dUTP and DY-555–dUTP into the nascent strand during synthesis. With our direct labeling protocol, the assay's runtime and handling steps could be reduced while the signal yield was increased. These features are very attractive for many detection formats but especially for point-of-care diagnostic kits that need to be simple enough to be performed by scientifically untrained personnel. KW - Method KW - Label KW - Fluorescence KW - RCA KW - Amplification KW - Oligonucleotide KW - DNA KW - Assay KW - Fluorophore KW - Dye content KW - Surface PY - 2012 DO - https://doi.org/10.1039/c2ay05760c SN - 1759-9660 SN - 1759-9679 VL - 4 IS - 5 SP - 1215 EP - 1220 PB - RSC Publ. CY - Cambridge AN - OPUS4-25839 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Linck, Lena A1 - Resch-Genger, Ute T1 - Identification of efficient fluorophores for the direct labeling of DNA via rolling circle amplification (RCA) polymerase phi29 JF - European journal of medicinal chemistry N2 - The enzymatic incorporation as well as the spectroscopic properties and photochemical stability of a series of fluorescent labels differing in dye class, charge, and rigidity were studied to identify new tools for signal enhancement in situ on microarrays without secondary labeling. These fluorophores were chosen to spectrally match or resemble the golden standard Cy3. With the rhodamine DY-555, that is three times more emissive than Cy3, we found a bright and stable chromophore, the spectroscopic properties of which are minimally influenced by dye microenvironment. KW - Rolling circle amplification (RCA) KW - DNA KW - Fluorescent label KW - Fluorescence quantum yield KW - Cyanine KW - Stability PY - 2010 DO - https://doi.org/10.1016/j.ejmech.2010.09.005 SN - 0009-4374 SN - 0223-5234 VL - 45 IS - 12 SP - 5561 EP - 5566 PB - EDIFOR CY - Paris AN - OPUS4-22470 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Resch-Genger, Ute A1 - Behnke, Thomas A1 - Brehm, Robert A1 - Grabolle, Markus A1 - Hennig, Andreas A1 - Hoffmann, Angelika A1 - Hoffmann, Katrin A1 - Linck, Lena A1 - Lochmann, Cornelia A1 - Pauli, Jutta A1 - Spieles, Monika A1 - Würth, Christian T1 - Funktionelle Chromophor-Systeme, innovative Validierungskonzepte und rückführbare Standards für die fluoreszenzbasierte multiparametrische Bioanalytik T2 - 5. Senftenberger Innovationsforum Multiparameteranalytik (Proceedings) N2 - 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. T2 - 5. Senftenberger Innovationsforum Multiparameteranalytik CY - Senftenberg, Deutschland DA - 10.03.2011 KW - Multiparametric KW - Multiplexing KW - Fluorescence KW - Nanoparticles KW - NIR dyes KW - Surface analysis KW - Quantum yield KW - Quantum dot KW - Lifetime PY - 2011 SP - 86 EP - 108 CY - Senftenberg AN - OPUS4-23635 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Linck, Lena A1 - Mayer-Enthart, Elke A1 - Sialelli, J. A1 - Rurack, Knut A1 - Köster, D. A1 - Seitz, H. A1 - Resch-Genger, Ute A1 - Panne, Ulrich T1 - Fluorescence Signal Amplification on DNA Microarrays via Rolling Circle Amplification (RCA) T2 - ANAKON 2009 T2 - ANAKON 2009 CY - Berlin, Germany DA - 2009-03-17 PY - 2009 AN - OPUS4-19131 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Linck, Lena A1 - Mayer-Enthart, Elke A1 - Sialelli, J. A1 - Rurack, Knut A1 - Köster, D. A1 - Seitz, H. A1 - Resch-Genger, Ute A1 - Panne, Ulrich T1 - Direct Enzymatic Incorporation of Fluorophores on DNA-Microarrays via Rolling Circle Amplification (RCA) T2 - Dechema Tagung T2 - Dechema Tagung CY - Frankfurt am Main, Germany DA - 2008-03-06 PY - 2008 AN - OPUS4-18548 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -