@article{IdaChanGloekleretal.2019, author = {Ida, Jeunice and Chan, Soo Khim and Gl{\"o}kler, J{\"o}rn and Lim, Yee Ying and Choong, Yee Siew and Lim, Theam Soon}, title = {G-Quadruplexes as An Alternative Recognition Element in Disease-Related Target Sensing}, series = {Molecules}, volume = {24}, journal = {Molecules}, number = {6}, issn = {1420-3049}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-11438}, year = {2019}, abstract = {G-quadruplexes are made up of guanine-rich RNA and DNA sequences capable of forming noncanonical nucleic acid secondary structures. The base-specific sterical configuration of G-quadruplexes allows the stacked G-tetrads to bind certain planar molecules like hemin (iron (III)-protoporphyrin IX) to regulate enzymatic-like functions such as peroxidase-mimicking activity, hence the use of the term DNAzyme/RNAzyme. This ability has been widely touted as a suitable substitute to conventional enzymatic reporter systems in diagnostics. This review will provide a brief overview of the G-quadruplex architecture as well as the many forms of reporter systems ranging from absorbance to luminescence readouts in various platforms. Furthermore, some challenges and improvements that have been introduced to improve the application of G-quadruplex in diagnostics will be highlighted. As the field of diagnostics has evolved to apply different detection systems, the need for alternative reporter systems such as G-quadruplexes is also paramount.}, language = {en} } @article{MorlockSubramanianZounietal.2023, author = {Morlock, Sascha and Subramanian, Senthil Kumar and Zouni, Athina and Lisdat, Fred}, title = {Closing the green gap of photosystem I with synthetic fluorophores for enhanced photocurrent generation in photobiocathodes}, series = {Chemical Science}, volume = {14}, journal = {Chemical Science}, publisher = {Royal Society of Chemistry (RSC)}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-17022}, pages = {1696 -- 1708}, year = {2023}, abstract = {One restriction for biohybrid photovoltaics is the limited conversion of green light by most natural photoactive components. The present study aims to fill the green gap of photosystem I (PSI) with covalently linked fluorophores, ATTO 590 and ATTO 532. Photobiocathodes are prepared by combining a 20 μm thick 3D indium tin oxide (ITO) structure with these constructs to enhance the photocurrent density compared to setups based on native PSI. To this end, two electron transfer mechanisms, with and without a mediator, are studied to evaluate differences in the behavior of the constructs. Wavelength-dependent measurements confirm the influence of the additional fluorophores on the photocurrent. The performance is significantly increased for all modifications compared to native PSI when cytochrome c is present as a redox-mediator. The photocurrent almost doubles from -32.5 to up to -60.9 μA cm-2. For mediator-less photobiocathodes, interestingly, drastic differences appear between the constructs made with various dyes. While the turnover frequency (TOF) is doubled to 10 e-/PSI/s for PSI-ATTO590 on the 3D ITO compared to the reference specimen, the photocurrents are slightly smaller since the PSI-ATTO590 coverage is low. In contrast, the PSI-ATTO532 construct performs exceptionally well. The TOF increases to 31 e-/PSI/s, and a photocurrent of -47.0 μA cm-2 is obtained. This current is a factor of 6 better than the reference made with native PSI in direct electron transfer mode and sets a new record for mediator-free photobioelectrodes combining 3D electrode structures and light-converting biocomponents.}, language = {en} } @article{MorlockSubramanianZounietal.2022, author = {Morlock, Sascha and Subramanian, Senthil Kumar and Zouni, Athina and Lisdat, Fred}, title = {Bio-inorganic hybrid structures for direct electron transfer to photosystem I in photobioelectrodes}, series = {Biosensors and Bioelectronics}, volume = {204}, journal = {Biosensors and Bioelectronics}, publisher = {Elsevier}, issn = {1873-4235}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-16770}, year = {2022}, abstract = {Synthetic materials can be combined with biological components in many ways. One example that provides scientists with multiple challenges is a photobioelectrode that converts sunlight into electrons in a biohybrid approach. In the present study several key parameters are evaluated concerning their influence on the direct electron transfer from a 3D indium tin oxide (ITO) electrode material to photosystem I (PSI) as a light-harvesting biomolecule. In contrast to previous investigations, no mediating molecule is added to shuttle the electrons to the luminal side of PSI. Thus, this setup is less complex than foregoing ones. The solution composition drastically influences the interaction of PSI with the ITO surface. Here, the application of higher buffer concentrations and the addition of salts are advantageous, whereas the nature of the buffer ions plays a minor role. The artificial electrode material's thickness is adjustable since a spin-coating procedure is used for preparation. With a 30 μm thick structure and immobilized PSI cathodic photocurrents up to 10.1 μA cm-2 are obtained at 100 mW cm-2 illumination intensity and an applied potential of -0.1V vs. Ag/AgCl. Over a period of three days the photobioelectrodes are illuminated for a total of 90 min and stored between the measurements at ambient temperature. The stability of the setup is noteworthy as still about 90\% of the photocurrent is retained. The photocathode described here offers many positive features, including a high onset potential for the photocurrent starting sligthly above the redox potentail of P700, and applicability in a wide pH range from pH 5 to 8.}, language = {en} } @article{GloeklerLimIdaetal.2021, author = {Gl{\"o}kler, J{\"o}rn and Lim, Theam Soon and Ida, Jeunice and Frohme, Marcus}, title = {Isothermal amplifications - a comprehensive review on current methods}, series = {Critical Reviews in Biochemistry and Molecular Biology}, journal = {Critical Reviews in Biochemistry and Molecular Biology}, publisher = {Taylor \& Francis}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-15154}, pages = {1 -- 44}, year = {2021}, abstract = {AbstractThe introduction of nucleic acid amplification techniques has revolutionized the field of medical diagnostics in the last decade. The advent of PCR catalyzed the increasing application of DNA, not just for molecular cloning but also for molecular based diagnostics. Since the introduction of PCR, a deeper understanding of molecular mechanisms and enzymes involved in DNA/RNA replication has spurred the development of novel methods devoid of temperature cycling. Isothermal amplification methods have since been introduced utilizing different mechanisms, enzymes, and conditions. The ease with which isothermal amplification methods have allowed nucleic acid amplification to be carried out has had a profound impact on the way molecular diagnostics are being designed after the turn of the millennium. With all the advantages isothermal amplification brings, the issues or complications surrounding each method are heterogeneous making it difficult to identify the best approach for an end-user. This review pays special attention to the various isothermal amplification methods by classifying them based on the mechanistic characteristics which include reaction formats, amplification information, promoter, strand break, and refolding mechanisms. We would also compare the efficiencies and usefulness of each method while highlighting the potential applications and detection methods involved. This review will serve as an overall outlook on the journey and development of isothermal amplification methods as a whole.}, language = {en} } @article{KallabisBeyerleinLisdat2024, author = {Kallabis, Conrad and Beyerlein, Peter and Lisdat, Fred}, title = {Quantitative determination of dopamine in the presence of interfering substances supported by machine learning tools}, series = {Bioelectrochemistry}, volume = {157}, journal = {Bioelectrochemistry}, number = {108667}, issn = {1567-5394}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-19277}, year = {2024}, abstract = {In the field of neuroscience as well as in the clinical setting, the neurotransmitter dopamine (DA) is an analyte which is important for research as well as medical purposes. There are plenty of methods available to measure dopamine quantitatively, with voltammetric ones such as differential pulse voltammetry (DPV) being among the most convenient and simple ones. However, dopamine often occurs, either naturally or because of the requirements of involved enzymatic systems, alongside substances that can influence the signal it produces upon electrochemical conversion. An example for such substances is the magnesium ion, which itself is not electrochemically active in the potential range needed for DA oxidation, but influences the dopamine signal. We have characterized the properties of DPV signals subject to the interaction between DA and Mg2+ and show that, although these properties are changing in a nonlinear fashion when both concentrations are varying, relatively simple linear mathematical models can be used to determine dopamine concentrations quantitatively in the presence of magnesium ions. The focus of this study is thus, the mathematical treatment of experimental data in order to overcome an analytical problem and not the investigation of the chemical background of DA-Mg2+ interaction.}, language = {en} } @article{GoebelMuellerTalkeetal.2024, author = {G{\"o}bel, Gero and M{\"u}ller, Florian and Talke, Anja and Ahnert, Uwe and Lisdat, Fred}, title = {Qualitative and quantitative protease activity tests based on protein degradation in three-dimensional structures}, series = {Bioelectrochemistry}, volume = {160}, journal = {Bioelectrochemistry}, issn = {1567-5394}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-19294}, year = {2024}, abstract = {The pattern of the activity of proteases is related to distinct physiological states of living organisms. Often activity changes of a certain protease can be assigned to a specific disease. Hence, they are useful biomarkers and a simple and fast determination method of their activity could be a valuable tool for the efficient monitoring of numerous diseases. Here, two different methods for the qualitative and quantitative determination of protease activity are demonstrated using the model system of proteinase K. The first test system is based on a protein-modified and colored 3D silica structure that changes color when exposed to the enzyme. This method has also been used for the detection of matrix metallo-protease 2 (MMP2) with gelatine as protease substrate on the plates. The second detection system uses the decrease in the voltammetric signal of a cytochrome c/DNA multilayer electrode after incubation with a protease to quantitatively determine its proteolytic activity. While activities down to 0.15 U/ml can be detected with the first method, the second one provides detection limits of about 0.03 U/ml (for proteinase K.) The functionality of both systems can be demonstrated and ways for further enhancement of sensitivity have been elucidated.}, language = {en} } @article{SuzukiHerkenhoffBroedeletal.2024, author = {Suzuki, Juliana Yumi and Herkenhoff, Marcos Edgar and Br{\"o}del, Oliver and Cucick, Ana Clara Candelaria and Frohme, Marcus and Saad, Susana Marta Isay}, title = {Exploring the potential of red pitaya pulp (Hylocererus sp.) as a plant-based matrix for probiotic delivery and effects on betacyanin content and flavoromics}, series = {Food Research International}, volume = {192}, journal = {Food Research International}, publisher = {Elsevier}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-19843}, year = {2024}, abstract = {This study evaluated the potential of red pitaya pulp fermented with Lacticaseibacillus paracasei subsp. paracasei F-19 (F-19) as a base for probiotic products. Physicochemical parameters, sugar, betacyanin, and phenolic contents, and antioxidant activity were analyzed over 28 days at 4 °C and compared to a non-fermented pulp, and to a pulp fermented with Bifidobacterium animalis subsp. lactis BB-12 (BB-12). Volatile compounds were identified using HS-SPME/GC-MS. Probiotic viability during storage and survival through in vitro-simulated gastrointestinal tract (GIT) stress were assessed. Red pitaya pulp, rich in moisture (85.83 g/100 g), carbohydrates (11.65 g/100 g), and fibers (2.49 g/100 g), supported fermentation by both strains. F-19 and BB-12 lowered pH, with F-19 showing stronger acidification, and maintained high viability (8.85-8.90 log CFU/mL). Fermentation altered sugar profiles and produced unique volatile compounds, enhancing aroma and sensory attributes. F-19 generated 2-phenylethanol, a unique flavor compound, absent in BB-12. Phenolic content initially increased but antioxidant activity decreased during storage. Betacyanin remained stable for up to 14 days. Red pitaya improved F-19 viability through the simulated GIT, while BB-12 populations significantly decreased (p < 0.05). These results suggest red pitaya pulp is a promising plant-based matrix for F-19, offering protection during digestion and highlighting its potential as a functional food with enhanced bioactive compound bioavailability and sensory attributes.}, language = {en} } @article{MorlockSubramanianZounietal.2021, author = {Morlock, Sascha and Subramanian, Senthil Kumar and Zouni, Athina and Lisdat, Fred}, title = {Scalable Three-Dimensional Photobioelectrodes Made of Reduced Graphene Oxide Combined with Photosystem I}, series = {ACS Applied Materials \& Interfaces}, volume = {13}, journal = {ACS Applied Materials \& Interfaces}, number = {9}, publisher = {American Chemical Society (ACS)}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-16760}, pages = {11237 -- 11246}, year = {2021}, abstract = {Photobioelectrodes represent one of the examples where artificial materials are combined with biological entities to undertake semi-artificial photosynthesis. Here, an approach is described that uses reduced graphene oxide (rGO) as an electrode material. This classical 2D material is used to construct a three-dimensional structure by a template-based approach combined with a simple spin-coating process during preparation. Inspired by this novel material and photosystem I (PSI), a biophotovoltaic electrode is being designed and investigated. Both direct electron transfer to PSI and mediated electron transfer via cytochrome c from horse heart as redox protein can be confirmed. Electrode preparation and protein immobilization have been optimized. The performance can be upscaled by adjusting the thickness of the 3D electrode using different numbers of spin-coating steps during preparation. Thus, photocurrents up to ∼14 μA/cm2 are measured for 12 spin-coated layers of rGO corresponding to a turnover frequency of 30 e- PSI-1 s-1 and external quantum efficiency (EQE) of 0.07\% at a thickness of about 15 μm. Operational stability has been analyzed for several days. Particularly, the performance at low illumination intensities is very promising (1.39 μA/cm2 at 0.1 mW/cm2 and -0.15 V vs Ag/AgCl; EQE 6.8\%).}, language = {en} } @misc{KeilRadkeTillichetal.2015, author = {Keil, Karolin and Radke, Lars and Tillich, Ulrich M. and Frohme, Marcus}, title = {Automatisierung des Bio-Plex Pro Analyseverfahrens}, series = {Wissenschaftliche Beitr{\"a}ge 2015}, volume = {19}, journal = {Wissenschaftliche Beitr{\"a}ge 2015}, issn = {0949-8214}, doi = {10.15771/0949-8214_2015_1_2}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-3524}, pages = {15 -- 19}, year = {2015}, abstract = {F{\"u}r den simultanen Nachweis mehrerer Analyten innerhalb einer Probe ist die Bead-basierte Multiplexanalytik ein h{\"a}ufig verwendetes Verfahren und wird beispielsweise zur Quantifizierung von Proteinen genutzt. Die aufwendige Durchf{\"u}hrung der Assays soll durch Automatisierung einerseits dem Anwender abgenommen werden, w{\"a}hrend andererseits die Genauigkeit und Reproduzierbarkeit der Analyse gesteigert wird. Die Automatisierung des Bio-Plex Pro™ Assays ist mit dem Pipettierroboter Tecan Freedom EVO 200 umgesetzt worden. Es ist ein Skript mit der Software Freedom EVOware® entwickelt worden, welches die Probenvorbereitung des Assays vollst{\"a}ndig {\"u}bernimmt. F{\"u}r einen Vergleich der manuellen und automatisierten Methode sind die humanen Zytokine IL-2, IL-4, IL-10, GM-CSF, IFN-γ und TNF-α in einer achtstufigen Standardverd{\"u}nnungsreihe und in unterschiedlich konzentrierten Proben gemessen worden. Die Berechnung der Streuungen (Standardabweichung und Variationskoeffizient) der einzelnen Standardverd{\"u}nnungsreihen sowie der Vergleich von gemessenen und erwarteten Konzentrationen der automatisierten und manuellen Methode zeigen, dass die Automatisierung neben der zeitlichen Optimierung auch die Genauigkeit und Reproduzierbarkeit der Analyse verbessert.}, language = {de} } @misc{BuetowPaenkeKafkaetal.2008, author = {B{\"u}tow, Sandra M. and P{\"a}nke, Oliver and Kafka, Jan and Lisdat, Fred}, title = {Goldchipelektroden zur elektrochemischen DNA-Detektion}, series = {Wissenschaftliche Beitr{\"a}ge 2008}, volume = {13}, journal = {Wissenschaftliche Beitr{\"a}ge 2008}, issn = {0949-8214}, doi = {10.15771/0949-8214_2008_1_4}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus-653}, pages = {25 -- 34}, year = {2008}, abstract = {Im folgenden Artikel werden einfache DNA-Sensoren vorgestellt, mit deren Hilfe es durch voltammetrische und impedimetrische Messmethoden m{\"o}glich ist, schnell, sensitiv und kosteng{\"u}nstig Einzelstrang-DNA (ssDNA) nachzuweisen. Beide Messprinzipien lassen neben der spezifischen Detektion auch die Quantifizierung von DNA-Sequenzen sowie den Nachweis von einzelnen Basenfehlpaarungen innerhalb dieser Sequenzen zu. F{\"a}nger- DNA wurde zu diesem Zweck mit dem 5'-Ende auf einer Goldoberfl{\"a}che immobilisiert. Die Hybridisierung mit einem Methylenblau (MB) markierten oder unmarkierten Probenstrang konnte dann mit Hilfe der Differenzpulsvoltammetrie DPV oder der elektrochemischen Impedanzspektroskopie nachgewiesen werden. Die voltammetrische Quantifizierung erfolgte in einem direkten und kompetitiven Ansatz, mit einem Detektionslimit von 30 nM bzw. 3 nM (bei Einsatz von 0,1 μM Kompetitor- DNA). Das Detektionslimit beim impedimetrischen Nachweis lag bei 100 nM DNA. Die hier vorgestellten Sensoren sind zum einen regenerierbar und k{\"o}nnen zum anderen {\"u}ber einen Zeitraum von zwei Monaten gelagert werden.}, language = {de} } @misc{RadkeLopezHemmerlingLubitzetal.2010, author = {Radke, Lars and L{\´o}pez-Hemmerling, Diana and Lubitz, Annika and Giese, Christoph and Wildenauer, Franz-Xaver and Frohme, Marcus}, title = {Etablierung verschiedener Bead-basierter Multiplexmethoden mit einem Suspensions Array-System f{\"u}r molekulardiagnostische Zwecke}, series = {Wissenschaftliche Beitr{\"a}ge 2009/2010}, volume = {14}, journal = {Wissenschaftliche Beitr{\"a}ge 2009/2010}, issn = {0949-8214}, doi = {10.15771/0949-8214_2010_1_1}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus-802}, pages = {6 -- 12}, year = {2010}, abstract = {Die simultane Bestimmung mehrerer Analyten und die Erstellung komplexer Parameterprofile erlangt immer gr{\"o}ßere Bedeutung in der heutigen Labordiagnostik. Die Bead-basierte Multiplexanalytik bietet hier eine flexible, schnelle und einfache Methode zur Erstellung individueller Analysen. Aufgrund der Vielzahl von Anwendungsm{\"o}glichkeiten, die diese moderne Nachweismethode im Bereich molekularbiologischer Fragestellungen bietet, ist die Etablierung der Beadbasierten Multiplexanalytik im Labor f{\"u}r Molekularbiologie und funktionelle Genomik der Technischen Hochschule Wildau von großem Nutzen. Zur Einarbeitung in das Testsystem wurden die Konzentrationen der Zytokine IL-2, IL-4, IFN-γ und TNF-α in Zellkultur{\"u}berst{\"a}nden mit kommerziellen Fertigsystemen gemessen und mit mRNA-Expressionsraten der gleichen Proben verglichen. Des Weiteren wurde ein Testsystem zum Nachweis von humanen Antik{\"o}rpern der Klassen IgG und IgM sowie deren antigen-spezifischer Anteil in Zellkultur{\"u}berst{\"a}nden entwickelt. Außerdem konnte durch die erfolgreiche Detektion von DNA-gekoppelten Beads mittels markierter Oligonukleotidsequenz die Kopplung und die Anwendbarkeit der Methode auf Bindungsexperimente mit Nukleotidsequenzen gezeigt werden.}, language = {de} } @misc{Fischer2013, author = {Fischer, Susanne}, title = {Experimentelle Therapieentwicklung f{\"u}r die akute lymphatische Leuk{\"a}mie}, series = {Wissenschaftliche Beitr{\"a}ge 2013}, volume = {17}, journal = {Wissenschaftliche Beitr{\"a}ge 2013}, issn = {0949-8214}, doi = {10.15771/0949-8214_2013_1_2}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-3146}, pages = {13 -- 16}, year = {2013}, abstract = {Bei Leuk{\"a}mie handelt es sich um eine Entartung von Vorl{\"a}uferzellen der weißen Blutzellen (Leukozyten). Die hier betrachtete Form dieser Erkrankung ist die akute lymphatische Leuk{\"a}mie(ALL ), bei der vor allem R{\"u}ckfallpatienten eine schlechte Diagnose haben. Demnach wird nach neuen Therapien gesucht, die neue Angriffspunkte f{\"u}r die Behandlung der ALL bieten. Die getestete Substanz 5-Azacytidin hat zwei therapeutische Wirkmechanismen. Zum einen das Hervorrufen des selbstinduzierte Zelltod (Apoptose) und zum anderen einen epigenetischen Effekt. Der epigenetische Effekt bewirkt eine Ver{\"a}nderung der Genregulation (Ver{\"a}nderung der Genaktivit{\"a}t) ohne die DNA - Sequenz zu beeinflussen. Dies kann mithilfe eines Screenings (Micoarraytechnologie) in behandelten und unbehandelten Proben verglichen werden. Da das Screening sehr global ist, erfolgt f{\"u}r eine Auswahl therapeutisch relevanter Gene eine Verifizierung mittels einer molekularbiologischen Methode (qRT -PCR ). Dabei wurden Gene mit erh{\"o}hter Aktivit{\"a}t best{\"a}tigt, MA EL; TM EM56; IL 1B; BCL 6; MS4A1 und FLT 3. F{\"u}r jedes dieser Gene konnte ein positiver Nutzen f{\"u}r die Therapie einer ALL diskutiert werden. F{\"u}r FLT 3 erfolgte sogar eine positive Kombination mit einer anderen therapeutischen Substanz. Der therapeutische Einsatz des 5-Azacytidins kann zuk{\"u}nftig eine wichtige Therapieoption f{\"u}r Patienten mit ALL darstellen.}, language = {de} } @misc{TanneGoebelLisdat2011, author = {Tanne, Christoph K. and G{\"o}bel, Gero and Lisdat, Fred}, title = {Entwicklung einer Glucosedehydrogenase-basierten Anode und deren Anwendung in einer Glucose/O2-Biobrennstoffzelle}, series = {Wissenschaftliche Beitr{\"a}ge 2011}, volume = {15}, journal = {Wissenschaftliche Beitr{\"a}ge 2011}, issn = {0949-8214}, doi = {10.15771/0949-8214_2011_1_2}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus-1039}, pages = {13 -- 21}, year = {2011}, abstract = {Unter Verwendung von mehrwandigen Kohlenstoffnanor{\"o}hren wurde in dieser Studie eine neuartige Anode zum Einsatz in Biobrennstoffzellen entwickelt. Dazu wurde das rekombinante Enzym Pyrrolochinolinchinon(PQQ)- abh{\"a}ngige Glucosedehydrogenase kovalent an eine aus PQQ bestehenden Zwischenschicht gekoppelt, welche zuvor an die Kohlenstoffnanor{\"o}hren adsorbiert war. Die Nanor{\"o}hren wurden aufgrund ihrer Thiolmodifikation chemisorptiv auf einer Goldelektrode gebunden. In glucosehaltiger L{\"o}sung konnte der Start eines katalytischen Stroms bei einem Potential von -80 mV vs. Ag/AgCl (1 MKCl) beobachtet werden. Unter Substrats{\"a}ttigung wurden Stromdichten im Bereich von 170 bis 200 μA/cm2 gemessen. Dieses System basiert auf einem mediatorvermittelten Elektronentransfer. Die entwickelte (PQQ)-GDH-MWCNT-Elektrode wurde mit einer MWCNT-modifizierten Elektrode kombiniert, bei der Bilirubinoxidase (BOD) als Biokatalysator fungiert. Daraus resultierte eine membranfreie Biobrennstoffzelle mit einem leichgewichtspotential von 600 mV und Leistungsdichten im Bereich von 20-25 μW/cm2.}, language = {de} } @article{FischbachLohBieretal.2017, author = {Fischbach, Jens and Loh, Qiuting and Bier, Frank Fabian and Lim, Theam Soon and Frohme, Marcus and Gl{\"o}kler, J{\"o}rn}, title = {Alizarin Red S for Online Pyrophosphate Detection Identified by a Rapid Screening Method}, series = {Scientific Reports}, volume = {7}, journal = {Scientific Reports}, issn = {2045-2322}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-9248}, year = {2017}, abstract = {We identified Alizarin Red S and other well known fluorescent dyes useful for the online detection of pyrophosphate in enzymatic assays, including the loop mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR) assays. An iterative screening was used for a selected set of compounds to first secure enzyme compatibility, evaluate inorganic pyrophosphate sensitivity in the presence of manganese as quencher and optimize conditions for an online detection. Of the selected dyes, the inexpensive alizarin red S was found to selectively detect pyrophosphate under LAMP and PCR conditions and is superior with respect to its defined red-shifted spectrum, long shelf life and low toxicity. In addition, the newly identified properties may also be useful in other enzymatic assays which do not generate nucleic acids but are based on inorganic pyrophosphate. Finally, we propose that our screening method may provide a blueprint for rapid screening of compounds for detecting inorganic pyrophosphate.}, language = {en} } @misc{RiedelKartchemnikSchoeningetal.2015, author = {Riedel, Marc and Kartchemnik, Julia and Sch{\"o}ning, Michael J. and Lisdat, Fred}, title = {Impedimetrischer DNA Nachweis - Schritte in Richtung sensorischer Anwendung}, series = {Wissenschaftliche Beitr{\"a}ge 2015}, volume = {19}, journal = {Wissenschaftliche Beitr{\"a}ge 2015}, issn = {0949-8214}, doi = {10.15771/0949-8214_2015_1_3}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:526-opus4-3538}, pages = {21 -- 28}, year = {2015}, abstract = {Diese Studie beschreibt einen labelfreien impedimetrischen Sensor auf der Grundlage von kurzen einzelstr{\"a}ngigen DNA-Erkennungselementen f{\"u}r den Nachweis von Hybridisierungsereignissen. Der Fokus der Arbeit liegt auf der Aufkl{\"a}rung des Einflusses der Ziel-DNA-L{\"a}nge und der Erkennungssequenzposition auf die sensorische Leistungsf{\"a}higkeit. Die impedimetrischen Messungen werden in Anwesenheit des Redoxsystems Kaliumhexacyanoferrat (II/III) durchgef{\"u}hrt und zeigen einen Anstieg des Durchtrittswiderstandes nach der Hybridisierung mit komplement{\"a}rer Ziel-DNA mit einer Nachweisgrenze im unteren nanomolaren Bereich. Nach der Hybridisierung kann die Regeneration des Sensors mit deionisiertem Wasser durch die Einstellung effektiver Konvektionsbedingungen erreicht werden und erm{\"o}glicht somit eine Wiederverwendbarkeit des Sensors. Untersuchungen zu l{\"a}ngeren Ziel-DNA-Str{\"a}ngen mit einem zur L{\"o}sung exponierten {\"U}berhang demonstrieren die Anwendbarkeit des impedimetrischen Nachweises f{\"u}r l{\"a}ngere Sequenzen. Allerdings resultiert eine zunehmende {\"U}berhangl{\"a}nge in einer verringerten Durchtrittswiderstands{\"a}nderung. Um die Impedanz{\"a}nderung f{\"u}r l{\"a}ngere Ziel-DNA zu erh{\"o}hen, wird die Erkennungssequenzposition ver{\"a}ndert, sodass ein kleiner {\"U}berhang zur Elektrode ausgerichtet ist. Die Ergebnisse legen nahe, dass DNA in direkter N{\"a}he zur Elektrode einen gr{\"o}ßeren Einfluss auf das impedimetrische Signal besitzt als weiter entfernte DNA.}, language = {de} }