Analytische Chemie
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Eingeladener Vortrag
- nein (287)
In this thesis a protocol based on a direct competitive and non-competitive immunoassay using the microarray format was developed to screen emulated hybridoma supernatants against the drug carbamazepine (CBZ). As support, epoxy slides were manufactured by coating microscope glass slides with epoxy silanes. The manufactured epoxy slides showed similar capacity for protein immobilization as commercially available NEXTERION slides, but showed a higher autofluorescence background. On the epoxy slides, protein A, Cys-A, G and Cys-G were immobilized as antibody capturing coating. It could be shown that the Cys-tag considerably increased the immobilization of all compared proteins, especially of protein G. Protein Cys-G immobilized at pH 8-9 was considered the most suitable protein for antibody immobilization due to good spot uniformity and a high binding capacity for IgG. Additionally, in immobilization experiments it could be shown, that 10-500 fold excess of bovine IgG, introduced by two different fetal bovine serum (FBS) products, showed no significant competition on the immobilization of mice or goat IgG on protein Cys-G. To examine the captured antibodies for affinity to CBZ, the CBZ-TOTA-DY654 tracer was custom synthesized. It could be demonstrated, that the tracer antibody interaction was competitive with the tracer precursor, CBZ-TOTA-NH2 and CBZ. Furthermore, the synthesized tracer was also successfully used by externals to evaluate CBZ affine hybriodoma cells in flow cytometry. In the simulated screening, five antibodies with known affinity to CBZ were diluted in a cell culture medium. The screening method allowed to differentiate between affine antibodies (IC50 < 20 µg/ml) and unaffine antibodies (IC50 > 20 µg/ml), when concentrations of 0.1 µg/ml IgG or greater have been used. Of five applied antibodies, all highly affine clones (CE2 and B3212M) were reported as positives while no false-positive samples were observed. The screening included competitive, with around 89500 fold excess of CBZ, and non-competitive tracer incubation on different segments of the same chip, which resulted in total signal suppression on the competitive segment. The screening was performed in 20 hours, only a few nanoliters, of each simulated supernatant, were consumed in the screening process, furthermore in the method standard 96 well MTPs were used and no cleanroom facilities were required.
Additive Manufacturing provides an important enabling technology for the digital transformation of the economy. As an emerging technology it has seen a remarkable development over the last three decades. Nevertheless, it is far from a broad adoption with several barriers to overcome yet. One of the major challenges is the lack of standards. The critical role of standardization for innovation is generally recognized, still the topic too often has been neglected in strategic roadmapping exercises for emerging technologies. Too little is known about the complex dynamics and interrelations of standardization and innovation. The anticipation of standardization needs and the timely and efficient implementation of standards is challenging. This paper aims at contributing to a better understanding of the role that standards play in the multi-dimensional system of innovation. It analyzes the trajectories of innovation in Additive Manufacturing in a systematic and holistic way, focusing on standardization activities with regard to coordination, stakeholders involved, the timing and types of standards developed. Putting standardization in context of the multi-dimensional innovation system of Additive Manufacturing the research shows where standards can support the diffusion of an emerging technology.
Leaking methane (CH4) from infrastructures, such as pipelines and landfills, is critical for the environment but can also pose a safety risk. To enable a fast detection and localization of these kind of leaks, we developed a novel robotic platform for aerial remote gas sensing. Spectroscopic measurement methods for remote sensing of selected gases lend themselves for use on mini-copters, which offer a number of advantages for inspection and surveillance over traditional methods. No direct contact with the target gas is needed and thus the influence of the aerial platform on the measured gas plume can be kept to a minimum. This allows to overcome one of the major issues with gas-sensitive mini-copters. On the other hand, remote gas sensors, most prominently Tunable Diode Laser Absorption Spectroscopy (TDLAS) sensors have been too bulky given the payload and energy restrictions of mini-copters. Here, we present the Unmanned Aerial Vehicle for Remote Gas Sensing (UAV-REGAS), which combines a novel lightweight TDLAS sensor with a 3-axis aerial stabilization gimbal for aiming on a versatile hexacopter. The proposed system can be deployed in scenarios that cannot be addressed by currently available robots and thus constitutes a significant step forward for the field of Mobile Robot Olfaction (MRO). It enables tomographic reconstruction of gas plumes and a localization of gas sources. We also present first results showing its performance under realistic conditions.
Isotope amount ratios (hereafter referred to as simply isotope ratios) are proving useful in an ever increasing array of applications that range from studies unravelling transport processes, to pinpointing the provenance of specific samples as well as trace element quantification by using isotope dilution mass spectrometry (IDMS). These expanding applications encompass fields as diverse as archaeology, food chemistry, forensic science, geochemistry, medicine and metrology. However, to be effective tools, the isotope ratio data must be reliable and traceable to enable the comparability of measurement.
The importance of traceability and comparability in isotope ratio analysis has already been recognized by the Inorganic Analysis Working Group (IAWG) within the CCQM. Three pilot studies have focused on the quality of isotope ratio determinations (P48 “U isotope ratios in urine”, P75 “stable isotopes in Methionine”, P105 “87Sr/86Sr in wine”). Moreover, isotope ratio measurements are fundamental to IDMS amount of substance determinations. For example, when Pb quantification using IDMS is undertaken, this requires the measurements of Pb isotope ratios. While the requirements for isotope ratio accuracy and precision in the case of of IDMS are generally quite modest, “absolute” Pb isotope ratio measurements for geochemical age dating and source rock characterization as well as forensic provenance and fingerprinting studies require Pb isotope ratio measurements of the highest quality. To support present and future CMCs on isotope ratio determinations, a Key Comparison was urgently needed. Therefore, it was decided at the IAWG meeting in Paris in April 2011 that a Key Comparison on the determination of Pb isotope ratios in a pure Pb solution and in a bronze sample should be organized and accompanied by a pilot study.
Measuring Pb isotope amount ratios in a pure Pb solution, while seemingly straight forward, rigorously tests the ability of analyst to correct for any instrumental effects (such as mass discrimination and blank correction) on the measured ratios. Pb, present in trace amounts in a metal matrix sample (e.g. Pb in bronze), provides a real world test of the whole chemical and instrumental procedure, from chemical separation and sample purification to analysis and subsequent correction of appropriate instrumental effects on the separated samples.
A suitable bronze material with a Pb mass fraction between 10 and 100 mg·kg-1 was available at BAM. A high purity solution of Pb with a mass fraction of approximately 100 mg·kg-1 was also available. By comparing the Pb isotope ratio results obtained for the bronze sample with the Pb isotope ratio results from the Pb solution, potential biases arising from the processing of the bronze sample could be effectively identified and separated from the instrumental effects arising from the measurement and data processing protocol.
The efficiency of gas sensor application for facilitating the safe use of hydrogen depends considerably on the sensor response to a change in hydrogen concentration. Therefore, the response time has been measured for five different-type commercially available hydrogen sensors. Experiments showed that all these sensors surpass the ISO 26142 standard; for the response times t90 values of 2 s to 16 s were estimated. Results can be fitted with an exponential or sigmoidal function. It can be demonstrated that the results on transient behaviour depend on both the operating parameters of sensors and Investigation methods, as well as on the experimental conditions: gas change rate and concentration jump.
Bacterial biofilms represent a ubiquitous form of microbial life on Earth. Due to an evolved armory of protean biological responses to external stimuli, bacteria are able to adhere to, colonize and thrive on virtually all surfaces, whether natural or synthetic, even in challenging environmental conditions. In addition to significant health risks, biofilms are among the salient contributors to the deterioration of metals and their alloys, thereby causing safety risks for technical equipment. Hence, understanding the interaction mechanisms of electroactive sessile bacteria with metal surfaces is vital for facilitating the development of efficient control strategies and novel anti-fouling surfaces in various industries and technologies.
The present study focusses on a combined spectroelectrochemical approach, melding methods of surface enhanced Raman spectroscopy (SERS) and electrochemical techniques, to investigate the chemical characteristics and redox activities of electroactive bacteria during the initial stages of biofilm formation. Gold has been selected as a model substrate due to its inert character, considerably high surface enhancement factor, as well as its capability to allow surface chemistry modifications and substrate polarization in order to precisely control the surface charge. Square wave voltammetry (SWV) and cyclic voltammetry (CV) studies have been performed for quantitative determination of flavin concentration and electrochemical impedance spectroscopy (EIS) has been utilized to study the changes in electrochemical processes within biofilms during different stages of growth. Shewanella sp. have been chosen as microorganisms within this work due to their versatile exoelectrogenic respiratory behavior and their distinct ability to reduce metals via extracellular electron transfer mechanisms involving self-secreted electron shuttle redox molecules such as flavins. To further explicate the process of diffusion of flavins within biofilms, a model system has been developed to simulate the structural features of the bacterial extracellular polymeric substances typically found in biofilms. This has been achieved by creating hydrogel films comprised of calcium-cross-linked alginate. The results demonstrate an interplay of factors contributing to the initial phases of bacterial settlement and biofilm formation as a function of environmental parameters. Furthermore, the results allow insight into the diffusion of flavins, much like they would in a natural biofilm, and how their redox behavior affects the biofilm development.
Capillary electrophoresis (CE) is an analytical technique that
separates analytes based on their electrophoretic mobility with
the use of an applied voltage.
Nanoparticles separation using CE exhibits good separation and
analysis efficiency, especially for nanoparticles smaller than 30
nm. The coupling of CE with inductively coupled plasma mass
spectrometry (ICP-MS) provides the possibility to detect
nanoparticles in the concentration range of nanograms per litre
and enables the simultaneous detection of nanoparticles and its
[1][2] ionic counter parts.
Our objective is to develop a fast and reliable separation method
for differently modified nanoparticles using capillary
electrophoresis.
The performance of a miniaturized channel for the separation of polymer and metal nanoparticles (NP) using Asymmetrical Flow Field-Flow Fractionation (AF4) was investigated and compared with a conventional AF4 system. To develop standard separation methods, experimental parameters like cross flow, gradient profile and injection time were varied and optimized.
Corresponding chromatographic parameters were calculated and compared. Our results indicate that the chromatographic resolution in the miniaturized channel is lower, whereas significantly shorter analyses time and less solvent consumption were obtained. Moreover, the limit of detection (LOD) and limit of quantification (LOQ) obtained from hyphenation with a UV-detector are obviously lower than in a conventional channel, which makes the miniaturized channel interesting for trace analysis.
A common method for nanoparticle separation, which was introduced in 1976 by Giddings, represents asymmetrical field-flow fractionation (AF4). It is a flow based separation method, which can be theoretically used to separate particles range from 1 nm to 50 µm. However, when the particles are smaller than 10 nm, separation with AF4 will become difficult to perform. Because in this case strong separation force, which induces aggregation of particles, should be applied. This will decrease recoveries of analytes and limit its application in accurate quantitative analysis. Capillary electrophoresis (CE) is another well-developed separation technique, in which samples will be separated in relation to their electrophoretic mobility. In recent years, CE has been used to separate different kinds of nanoparticles like, gold colloids or CdSe Quantum dots. However, till now only separation of particles smaller than 50 nm was reported. Because large size distribution of bigger particles will result in strong peak broadening and long separation time.
A two-dimensional coupling of AF4 and CE might provide us a new separation method, which can extend the separation ranges of both methods and be a way to characterise particles with large size distributions.
Inhibition of Herpes Virus by Specific and Non-specific Interactions With Graphene Conjugates
(2017)
Herpes viruses (HSV) are global, host-adapted pathogens that cause a widespread diversity of diseases. The frequency of HSV infections all over the world has amplified over the last years, making it a major concern in the area of public health. Therefore, synthesis of systems that can inhibit development of these viruses is required. Various compounds already have shown inhibition of HSV, but concentration of these inhibitors is relatively high and resistance against those drugs is challenging.
Combination of biological knowledge, about structure of the active site on the surface of HSV that is responsible for inhibition of the pathogen, with the chemistry of graphene results in 2D systems with the ability of specific and nonspecific interactions with HSV. In this work, 2D nanomaterials with picomolar IC50 against HSV are synthesized by conjugation of peptides to the surface of graphene.
2D nanomaterials are characterized by various methods, including XPS, AFM and IR. Biological evaluation showed high potency of synthesized nanomaterials to inhibit HSV and therefore underlined possibility to use such materials in future biomedical applications.