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Therapeutische Antikörper sind innerhalb weniger Jahre zur wichtigsten pharmazeutischen Produktklasse aufgestiegen. Für 2023 werden weltweite Umsätze von über 200 Milliarden USD erwartet. Auch diagnostische Antikörper sind mittlerweile unverzichtbare Produkte, auf deren Basis zahllose Immunoassays und andere Schnelltests entwickelt wurden. Neben der bereits sehr aufwendigen Herstellung von Antikörpern ist deren Aufreinigung aus komplexen Zellkulturmedien oder Blutseren und -plasmen zu einem Engpass in der Produktion und Nutzung dieser komplexen Proteine geworden. Schnelle und einfache Reinigungsmethoden für Antikörper sind daher sehr gefragt.
The primary screening of hybridoma cells is a time-critical and laborious step during the development of monoclonal antibodies. Often critical errors occur in this phase, which supports the notion that the generation of monoclonal antibodies with hybridoma technology is difficult to control and hence a risky venture. We think that it is crucial to improve the screening process to eliminate most of the immanent deficits of the conventional approach. With this new microarray-based procedure, several advances could be achieved: Selectivity for excellent binders, high throughput, reproducible signals, avoidance of misleading avidity (multivalency) effects, and simultaneous performance of competition experiments. The latter can directly be used to select clones of desired cross-reactivity properties. In this paper, a model system with two excellent clones against carbamazepine, two weak clones and blank supernatant has been designed to examine the effectiveness of the new system. The excellent clones could be detected largely independent of the IgG concentration, which is unknown during the clone screening since the determination and subsequent adjustment of the antibody concentration is not possible in most cases. Furthermore, in this approach, the enrichment, isolation, and purification of IgG for characterization is not necessary. Raw cell culture supernatant can be used directly, even when fetal calf serum (FCS) or other complex media had been used. In addition, an improved method for the oriented antibody-immobilization on epoxy-silanized slides is presented. Based on the results of this model system, we conclude that this approach should be preferable to most other protocols leading to many of false positives, causing expensive and lengthy confirmation steps to weed out the poor clones.
Solutions of organic analytes of known mass fraction are typically used to calibrate the measurement processes used to determine these compounds in matrix samples. Appropriate value assignments and uncertainty calculations for calibration solutions are critical for accurate measurements. Evidence of successful participation in formal, relevant international comparisons is needed to document measurement capability claims (CMCs) made by national metrology institutes (NMIs) and designated institutes (DIs). To enable NMIs and DIs to update or establish their claims, in 2015 the Organic Analysis Working Group (OAWG) sponsored CCQM-K131 "Low-Polarity Analytes in a Multicomponent Organic Solution: Polycyclic Aromatic Hydrocarbons (PAHs) in Acetonitrile".
Polycyclic aromatic hydrocarbons (PAHs) result from combustion sources and are ubiquitous in environmental samples. The PAH congeners, benz[a]anthracene (BaA), benzo[a]pyrene (BaP), and naphthalene (Nap) were selected as the target analytes for CCQM-K131. These targets span the volatility range of PAHs found in environmental samples and include potentially problematic chromatographic separations. Nineteen NMIs participated in CCQM-K131. The consensus summary mass fractions for the three PAHs are in the range of (5 to 25) μg/g with relative standard deviations of (2.5 to 3.5) %.
Successful participation in CCQM-K131 demonstrates the following measurement capabilities in determining mass fraction of organic compounds of moderate to insignificant volatility, molar mass of 100 g/mol up to 500 g/mol, and polarity pKow < −2 in a multicomponent organic solution ranging in mass fraction from 100 ng/g to 100 μg/g: (1) value assignment of primary reference standards (if in-house purity assessment carried out), (2) value assignment of single and/or multi-component organic solutions, and (3) separation and quantification using gas chromatography or liquid chromatography.
CRM (Certified Reference Material) BAM-U116/CGL306 “Cyanide in soil” was produced within a framework of cooperation between CGL (Central Geological Laboratory) of Mongolia and Federal Institute for Materials Research and Testing (BAM) of Germany in 2013-2017. The CRM BAM-U116/CGL306 represents a mixture of a sandy soil collected from a contaminated former gasworks area in the Berlin region (Germany) and an unpolluted sandy soil from Nalaikh region (Mongolia).
The bulk candidate material for this reference material was prepared at CGL CRM Laboratory exclusively destined to the preparation of reference materials and equipped with modern technical equipment. Homogeneity, stability and shelf life were studied in full compliance with ISO Guide 35. The CRM was evaluated as sufficiently homogeneous. Statistical evaluation of certification analysis
was software packages SoftCRM and ProLab Plus. Certified value of total cyanide of the CRM is 12.0 mg/kg and expanded uncertainty was assigned as 0.8 mg/kg. The intended purpose of this material is the verification of analytical results obtained for the mass fraction of total cyanide in soils and soil-like materials applying the standardized procedure ISO 11262:2011. As any reference material, it can also be used for routine performance checks (quality control charts) or validation studies.
The trafficking of illegal drugs by criminal networks at borders, harbors, or airports is an increasing issue in public health as these routes ensure the main supply of illegal drugs. The prevention of drug smuggling, including the installation of scanners and other analytical devices to detect ultra-small traces of drugs within a reasonable time frame, remains a challenge. The presented immunosensor is based on a monolithic affinity column with a large excess of immobilized hapten, which traps fluorescently labeled antibodies as long as the analyte cocaine is absent. In the presence of the drug, some binding sites of the antibody will be blocked, which leads to an immediate breakthrough of the labeled protein, detectable by highly sensitive laser-induced fluorescence with the help of a Peltier-cooled complementary metal-oxide-semiconductor (CMOS) camera. Liquid handling is performed with high-precision syringe pumps and microfluidic chip-based mixing devices and flow cells. The biosensor achieved limits of detection of 23 pM (7 ppt) of cocaine with a response time of 90 seconds and a total assay time below 3 minutes. With surface wipe sampling, the biosensor was able to detect 300 pg of cocaine. This immunosensor belongs to the most sensitive and fastest detectors for cocaine and offers near-continuous analyte measurement.
Near-infrared (NIR) spectroscopy is a promising candidate for low-cost, nondestructive, and highthroughput mass quantification of microplastics in environmental samples. Widespread application of the technique is currently hampered mainly by the low sensitivity of NIR spectroscopy compared to thermoanalytical approaches commonly used for this type of analysis. This study shows how the application of NIR spectroscopy for mass quantification of microplastics can be extended to smaller analyte levels by combining it with a simple and rapid microplastic enrichment protocol. For this purpose, the widely used flotation of microplastics in a NaCl solution, accelerated by centrifugation, was chosen which allowed to remove up to 99 % of the matrix at recovery rates of 83–104 %. The spectroscopic measurements took place directly on the stainless-steel filters used to collect the extracted particles to reduce sample handling to a minimum. Partial least squares regression (PLSR) models were used to identify and quantify the extracted microplastics in the mass range of 1–10 mg.
The simple and fast extraction procedure was systematically optimized to meet the requirements for the quantification of microplastics from common PE-, PP-, and PS-based packaging materials with a particle size < 1 mm found in compost or soils with high natural organic matter content (> 10 % determined by loss on ignition). Microplastics could be detected in model samples at a mass fraction of 1 mg g-1. The detectable microplastic mass fraction is about an order of magnitude lower compared to previous studies using NIR spectroscopy without additional enrichment. To emphasize the cost-effectiveness of the method, it was implemented using some of the cheapest and most
compact NIR spectrometers available.
2,4-D ist ein in der Landwirtschaft weitverbreitetes Pflanzenschutzmittel, das Grundwasser kontaminiert, sich innerhalb der Nahrungskette anreichert und Umwelt- und Gesundheitsprobleme verursachen kann. Hier stellen die Autoren ein mikrofluidisches Nachweissystem für die Echtzeitdetektion von 2,4-D in Grund- oder Oberflächenwasser vor. Es basiert auf der Kombination 2,4-D-selektiver, fluoreszierender, molekular geprägter Polymer-(MIP-)Mikropartikel mit einem 3D-mikrofluidischen Extraktions- und Detektionssystem. Messungen vor Ort sollen damit künftig möglich sein.
Reported here is the production of molecularly imprinted polymer (MIP) films, integrating a fluorescent moiety that serves as both an element for template interaction and signalling, for the thermal and optical detection of the beta-lactam antibiotic nafcillin. Fluorescein methacrylate (FluMa) was synthesized and introduced during the molecular imprinting process as the sole monomer and in a 1 : 1 mixture with methacrylic acid (MAA), allowing to draw first conclusions on the MIP formation potential of such a rather large and rigid monomer. At first, MIP microparticles containing FluMa were prepared by free radical polymerisation. Optical batch rebinding experiments revealed that FluMa can act as a functional monomer for selective detection of nafcillin; however, the addition of MAA as co-monomer significantly improved performance. Subsequently, thin MIP films containing FluMa were deposited onto functionalised glass slides and the influence of porogen, drying time, and monomer composition was studied. These MIP-functionalised glass electrodes were mounted into a customised 3D-printed flow cell, where changes in the liquid were either evaluated with a thermal device or using fluorescence bright field microscopy. Thermal analysis demonstrated that multiple MIP layers enhanced sensor specificity, with detection in the environmentally relevant range. The fluorescence bright field microscope investigations validated these results, showing an increase in the fluorescence intensity upon exposure of the MIP-functionalised glass slides to nafcillin solutions. These are promising results for developing a portable sensor device that can be deployed for antibiotics outside of a dedicated laboratory environment, especially if sensor design and fluorophore architecture are optimised.