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Development of a Genus-Specific Antigen Capture ELISA for Orthopoxviruses – Target Selection and Optimized Screening
No full textOrthopoxvirus species like cowpox, vaccinia and monkeypox virus cause zoonotic infections in humans worldwide. Infections often occur in rural areas lacking proper diagnostic infrastructure as exemplified by monkeypox, which is endemic in Western and Central Africa. While PCR detection requires demanding equipment and is restricted to genome detection, the evidence of virus particles can complement or replace PCR. Therefore, an easily distributable and manageable antigen capture enzyme-linked immunosorbent assay (ELISA) for the detection of orthopoxviruses was developed to facilitate particle detection. By comparing the virus particle binding properties of polyclonal antibodies developed against surface-exposed attachment or fusion proteins, the surface protein A27 was found to be a well-bound, highly immunogenic and exposed target for antibodies aiming at virus particle detection. Subsequently, eight monoclonal anti-A27 antibodies were generated and characterized by peptide epitope mapping and surface plasmon resonance measurements. All antibodies were found to bind with high affinity to two epitopes at the heparin binding site of A27, toward either the N- or C-terminal of the crucial KKEP-segment of A27. Two antibodies recognizing different epitopes were implemented in an antigen capture ELISA. Validation showed robust detection of virus particles from 11 different orthopoxvirus isolates pathogenic to humans, with the exception of MVA, which is apathogenic to humans. Most orthopoxviruses could be detected reliably for viral loads above 1 × 103 PFU/mL. To our knowledge, this is the first solely monoclonal and therefore reproducible antibody-based antigen capture ELISA able to detect all human pathogenic orthopoxviruses including monkeypox virus, except variola virus which was not included. Therefore, the newly developed antibody-based assay represents important progress towards feasible particle detection of this important genus of viruses
IEA SHC Task 42 / ECES Annex 29 WG A1: Engineering and Processing of PCMs, TCMs and Sorption Materials
No full textAn overview on the recent results on the engineering and characterization of sorption materials, PCMs and TCMs investigated in the working group WG A1 “Engineering and processing of TES materials” of IEA SHC Task 42 / ECES Annex 29 (Task 4229) entitled “Compact Thermal Energy Storage” is presented
Evaluation of Fabry-Perot polymer film sensors made using hard dielectric mirror deposition
No full textFabry-Perot (FP) polymer film sensors offer high acoustic sensitivity, small element sizes, broadband frequency response and optical transmission to enable high resolution, backward mode photoacoustic (PA) imaging. Typical approaches to sensor fabrication involve the deposition of stacks of alternating dielectric materials to form interferometer mirrors, which are separated by a polymer spacer. If hygroscopic soft dielectric materials are used, a protective polymer layer is typically required. In this study, methods for the deposition of water-resistant, hard dielectric materials onto polymers were explored to improve the robustness and performance of the sensors. This involved the optimisation of the fabrication process, the optical and acoustic characterisation of the sensors, and a comparison of the frequency response with the output of an acoustic forward model. The mirrors, which were separated by a 20 μm Parylene spacer, consisted of eight double layers of Ta2O5 and SiO2 deposited onto polymer substrates using temperature-optimised electron vapour deposition. The free spectral range of the interferometer was 32 nm, its finesse FR = 91, and its visibility V = 0.72. The noise-equivalent pressure was 0.3 kPa (20 MHz bandwidth). The measured frequency response was found to be more resonant at 25 MHz compared to sensors with soft dielectric mirrors, which was also in good agreement with the output of a forward model of the sensor. The sensors were used in a PA scanner to acquire 3-D images in tissue phantoms
Towards a novel bioelectrocatalytic platform based on “wiring” of pyrroloquinoline quinone-dependent glucose dehydrogenase with an electrospun conductive polymeric fiber architecture
No full textElectrospinning is known as a fabrication technique for electrode architectures that serve as immobilization matrices for biomolecules. The current work demonstrates a novel approach to construct a conductive polymeric platform, capable not only of immobilization, but also of electrical connection of the biomolecule with the electrode. It is produced upon electrospinning from mixtures of three different highly conductive sulfonated polyanilines and polyacrylonitrile on ITO electrodes. The resulting fiber mats are with a well-retained conductivity. After coupling the enzyme pyrroloquinoline quinone-dependent glucose dehydrogenase (PQQ-GDH) to polymeric structures and addition of the substrate glucose an efficient bioelectrocatalysis is demonstrated. Depending on the choice of the sulfonated polyanilline mediatorless bioelectrocatalysis starts at low potentials; no large overpotential is needed to drive the reaction. Thus, the electrospun conductive immobilization matrix acts here as a transducing element, representing a promising strategy to use 3D polymeric scaffolds as wiring agents for active enzymes. In addition, the mild and well reproducible fabrication process and the active role of the polymer film in withdrawing electrons from the reduced PQQ-GDH lead to a system with high stability. This could provide access to a larger group of enzymes for bioelectrochemical applications including biosensors and biofuel cells
A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies
No full textThe aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale
Historical Prospective: Boltzmann’s versus Planck’s State Counting—Why Boltzmann Did Not Arrive at Planck’s Distribution Law
No full textWhy does Planck (1900), referring to Boltzmann’s 1877 probabilistic treatment, obtain his quantum distribution function while Boltzmann did not? To answer this question, both treatments are compared on the basis of Boltzmann’s 1868 three-level scheme (configuration—occupation—occupancy). Some calculations by Planck (1900, 1901, and 1913) and Einstein (1907) are also sketched. For obtaining a quantum distribution, it is crucial to stick with a discrete energy spectrum and to make the limit transitions to infinity at the right place. For correct state counting, the concept of interchangeability of particles is superior to that of indistinguishability
Untersuchung zum Ausfall der Strangführungsrollen in der horizontalen Zone einer Brammenstranggussanlage
No full textEs besteht ein Problem durch den häufigen Ausfall der Strangführungsrollen und die resultierenden erheblichen Verluste durch den Produktionsausfall, welcher beim Ersetzen der Rolleneinheiten entsteht.
Um die Ursachen zu ergründen wurden, zunächst die Randbedingungen und das Schadensbild analysiert. Daraus entstand die Hypothese, dass allgemein eine zu hohe Temperaturbelastung für die Schädigung der Lagerkomponenten verantwortlich ist und dies die grundlegende Ursache für die weitere Zerstörung darstellt.
Folglich musste eine Messeinrichtung installiert werden, um genauere Erkenntnisse über den Überhitzungsvorgang zu erlangen. Zu diesem Zweck wurden mehrere Verfahren zur Temperaturmessung miteinander verglichen, woraufhin eine Auswahl anhand einer Nutzwertanalyse erfolgte.
Die Auswertung der gewonnenen Messdaten zeigte, dass die Charakteristik der aufgezeichneten Überhitzungen sehr variabel ist. Weiterhin konnte keine Abhängigkeit von einem Prozessparameter ermittelt werden. Die genauer betrachteten Vorgänge konnten nicht oder nur eingeschränkt als Ursache für die Überhitzung nachgewiesen werden.
Wenngleich somit weitere Untersuchungen erforderlich sind, wurden mögliche Maßnahmen zur Lösung des Problems vorgeschlagen.The problem deals with the frequent malfunction of the guide rolls and the following serious loss through the lost output, which develops because of the replacement of the rolls.
First of all, the boundary conditions and the failure mode were analyzed to fathom the possible causes. Hence, the assumption arose that the general reason for the damage of the bearing components is that the temperature stress is too high, which is the reason for the further destruction. Consequently, a system of measurements has to be installed to get detailed insights about overheating. For this purpose, different methods of temperature detections were compared, whereupon a choice occurred on the basis of a utility analysis.
The evaluation of the achieved measurement data showed that the characteristic of the recorded overheating is very different. Furthermore, a dependence on a process parameter cannot be proved. It was not possible, or just restricted, to verify the eyed instances as a cause of the high temperature.
Although more researches are necessary, possible actions to solve the problem were proposed
Wo bin ich? iBeacons im Einsatz in der Bibliothek der TH Wildau
Get PDFMit diesem Beitrag soll der Einsatz von iBeacons als Basis eines Indoor-Ortungssystems in der Hochschulbibliothek der Technischen Hochschule Wildau vorgestellt werden. Für die Ortung innerhalb von Gebäuden sind bisher unterschiedliche Strategien zum Einsatz gekommen. Die von Apple Inc. entwickelte iBeacon-Technologie kommuniziert über Bluetooth Low Energy (BLE) und setzt somit eine Technologie ein, die einen großen Kreis von Nutzern erreicht. Als wesentlicher Teil der Campus-App Unidos Wildau operiert das Ortungssystem auf der Basis des Kartendienstes iCampus Wildau Maps. Gemachte Erfahrungen mit der iBeacon-Technologie werden ebenso im Fokus dieses Beitrags stehen wie Ergebnisse der Implementierung unseres Ortungssystems sowie Pläne für die Weiterentwicklung.This article describes the usage of the iBeacon technology as the basis for an indoor positioning system within the library of the University of Applied Sciences Wildau. Developed by Apple Inc., this technology is based on the widespread Bluetooth Low Energy (BLE) standard, thus allowing the implementation reach a large number of client devices. The article will focus on the gained experiences so far and on the results of the implementation of a positioning algorithm within our iOS and Android apps, as well as the use of our map service, iCampus Wildau Maps, as the interface for displaying the user’s position. It will briefly discuss possible enhancements and present future plans for further development
The use of fluorescence microscopy and image analysis for rapid detection of non-producing revertant cells of Synechocystis sp. PCC6803 and Synechococcus sp. PCC7002
No full textEthanol production via genetically engineered cyanobacteria is a promising solution for the production of biofuels. Through the introduction of a pyruvate decarboxylase and alcohol dehydrogenase direct ethanol production becomes possible within the cells. However, during cultivation genetic instability can lead to mutations and thus loss of ethanol production. Cells then revert back to the wild type phenotype.
A method for a rapid and simple detection of these non-producing revertant cells in an ethanol producing cell population is an important quality control measure in order to predict genetic stability and the longevity of a producing culture. Several comparable cultivation experiments revealed a difference in the pigmentation for non-producing and producing cells: the accessory pigment phycocyanin (PC) is reduced in case of the ethanol producer, resulting in a yellowish appearance of the culture. Microarray and western blot studies of Synechocystis sp. PCC6803 and Synechococcus sp. PCC7002 confirmed this PC reduction on the level of RNA and protein
Adaptive Evolution und Screening bei Cyanobakterien - Erzeugung und Untersuchung von Thermotoleranz in Synechocystis sp. PCC 6803
No full textZiel dieser Arbeit war die Erhöhung der Temperaturtoleranz des Cyanobakteriums Synechocystis sp. PCC 6803 mittels ungerichteter Mutagenese und adaptiver Evolution. Trotz des erneuten Interesses an Cyanobakterien und Mikroalgen in den letzten Jahren, gibt es nur relativ wenige aktuelle Studien zum Einsatz dieser Methoden an Cyanobakterien. Zur Analyse eines mittels Mutagenese erzeugten Gemischs an Stämmen, ist es von großem Vorteil Hochdurchsatz-Methoden zur Kultivierung und zum Screening einsetzen zu können. Auf Basis eines Pipettierroboters wurde solch eine Plattform für phototrophe Mikroorganismen neu entwickelt und folgend stetig verbessert. Die Kultivierung erfolgt in 2,2ml Deepwell-Mikrotiterplatten innerhalb einer speziell angefertigten Kultivierungskammer. Schüttelbedingungen, Beleuchtung, Temperatur und CO2-Atmosphäre sind hierbei vollständig einstellbar.Die Plattform erlaubt semi-kontinuierliche Kultivierungen mit automatisierten Verdünnungen von hunderten Kulturen gleichzeitig. Automatisierte Messungen des Wachstums, des Absorptionsspektrums, der Chlorophyllkonzentration, MALDI-TOF-MS sowie eines neu entwickelten Vitalitätsassays wurden etabliert. Für die Mutagenese wurden die Letalität- und die nicht-letale Punktmutationsrate von ultravioletter Strahlung und Methylmethansulfonat für Synechocystis charakterisiert. Synechocystis wurde mit den so ermittelten optimalen Dosen mehrfach behandelt und anschließend einer in vivo Selektion unterzogen. Somit wurde dessen Temperaturtoleranz um bis zu 3°C erhöht. Über die Screeningplattform wurden die thermotolerantesten monoklonalen Stämme identifiziert. Nach einer Validierung wurde das vollständige Genom der Stämme sequenziert. Hierdurch wurden erstmals Mutationen in verschiedenen Genen mit der Langzeittemperaturtoleranz von Synechocystis in Verbindung gebracht. Bei einigen dieser Gene ist es sehr unwahrscheinlich, dass sie mittels anderer Verfahren hätten identifiziert werden können.The goal of this work was the increase of the thermal tolerance of the cyanobacteria Synechocystis sp. PCC 6803 via random mutagenesis and adaptive evolution. Even with the renewed interest in cyanobacteria in the recent years, there is relatively limited current research available on the application of these methods on cyanobacteria. To analyse a mixture of various strains typically obtained through random mutagenesis, a method allowing high-throughput miniaturized cultivation and screening is of great advantage. Based on a pipetting robot a novel high-throughput screening system suitable for phototrophic microorganisms was developed and then constantly improved. The cultivation was performed in 2,2 ml deepwell microtiter plates within a cultivation chamber outfitted with programmable shaking conditions, variable illumination, variable temperature, and an adjustable CO2 atmosphere. The platform allows semi-continuous cultivation of hundreds of cultures in parallel. Automated measurements of growth, full absorption spectrum, chlorophyll concentration, MALDI-TOF-MS, as well as a novel vitality measurement protocol, have been established. Prior to the mutagenesis, the lethality and rate of non-lethal point mutations of ultraviolet radiation and methyl-methanesulphonate were characterized for Synechocystis. The thus determined optimal dosages were applied to Synechocystis followed by in vivo selection in four rounds of mutagenesis, thereby raising its temperature tolerance by 3°C. The screening platform was used to identify the most thermotolerant monoclonal strains. After validation, their whole genomes were sequenced. Thus mutations in various genes were identified which promote the strains'' thermal tolerance. For some of the genes it is very unlikely that their link to high thermal tolerance could have been identified by other approaches