Analytische Chemie
Filtern
Erscheinungsjahr
- 2019 (40) (entfernen)
Dokumenttyp
- Posterpräsentation (15)
- Zeitschriftenartikel (14)
- Vortrag (11)
Sprache
- Englisch (40)
Schlagworte
- Fluorescence (40) (entfernen)
Organisationseinheit der BAM
- 1 Analytische Chemie; Referenzmaterialien (37)
- 1.2 Biophotonik (24)
- 1.9 Chemische und optische Sensorik (11)
- 6 Materialchemie (4)
- 6.3 Strukturanalytik (3)
- 1.0 Abteilungsleitung und andere (1)
- 1.1 Anorganische Spurenanalytik (1)
- 1.5 Proteinanalytik (1)
- 6.5 Synthese und Streuverfahren nanostrukturierter Materialien (1)
- 8 Zerstörungsfreie Prüfung (1)
- 8.1 Sensorik, mess- und prüftechnische Verfahren (1)
Eingeladener Vortrag
- nein (11)
A “turn-on” fluorescence sensing system based on a BODIPY-cobaloxime complex for the detection of H2S in liquid and gas phase was developed. To that aim, two cobaloxime complexes bearing an axial pyridyl-BODIPY ligand were initially evaluated as sensitive fluorescent HS− indicators in aqueous solution. The sensing mechanism involves the selective substitution of the BODIPY ligand by the HS− anion at the cobalt center, which is accompanied by a strong fluorescence enhancement. The selection of a complex with an ideal stability and reactivity profile toward HS− relied on the optimal interaction between the cobalt metal-center and two different pyridyl BODIPY ligands. Loading the best performing BODIPY-cobaloxime complex onto a polymeric hydrogel membrane allowed us to study the selectivity of the probe for HS− against different anions and cysteine. Successful detection of H2S by the fluorescent “light-up” membrane was not only accomplished for surface water but could also be demonstrated for relevant H2S concentrations in gas phase.
Chlorination of pool water and wastewater, in food and pharmaceutical production, as well as in pesticide and paper manufacturing is a routinely used technique. However, the amount of chlorine in water must be strictly adjusted, to ensure enough concentration to kill pathogenic bacteria and viruses, while preventing too high concentrations inducing negative effects on human health. As an indicator, a molecular fluorescent probe based on a BODIPY structure was designed. This indicator exhibits a sensitive and selective fluorescence response upon increasing concentrations of hypochlorite in aqueous solvent mixtures. Real-time analyses became possible after the integration of this fluorescent indicator into newly designed 2D & 3D microfluidic chips incorporating a passive sinusoidal mixer and a micro-hydrocyclone, respectively. A comparison of the two microfluidic systems, including their ability to prevent accumulation or circulation of microbubbles, has shown excellent fluidic behaviour for the micro-hydrocyclone device. This system was distinctly more robust against gas bubbles, showed a higher signal gain and allowed to halve the limit of detection to 0.02 mg L–1. The use of the 3D system to quantify the chlorine content of pool water samples for sensitive and quantitative chlorine monitoring has been demonstrated.
The reliable identification and quantification of phosphorylated amino acids, peptides and proteins is one of the key challenges in contemporary bioanalytical research, noteworthy, to diagnose and treat diseases at an early developmental stage. Miniaturised sensing devices like microfluidic chips combined with “smart” detection chemistry, simple data assessment, processing and presentation are very attractive for benchtop use in clinical environments.
We developed novel synthetic probes targeting phosphorylated amino acids, based on core-shell microparticles consisting of a silica core coated with a molecularly imprinted polymer (MIP) shell. These “plastic antibodies” are extremely robust, resist denaturing solvents and elevated temperatures, can be reproducibly produced at low cost, and potentially overcome many of the practical problems in current bioanalytical detection strategies. The MIP layer contains a fluorescent probe monomer, binds selectively to phosphorylated tyrosine (pTyr) with a significant imprinting factor higher than 3.5 and responds with a “lighting-up” of its fluorescence accompanied by the development of a strongly red-shifted emission band toward the analyte.
In analogy to our previous work [4], the bead-based ratiometric detection scheme has also been successfully transferred to a microfluidic chip format to demonstrate its applicability to rapid assays. Such a miniaturised device could yield an automated pTyr measurement system in the future. The setup was built by coupling a modular microfluidic system [5] for amino acid functionalisation (Fmoc protection) and a multi-layer PDMS/Teflon/glass microfluidic chip [6] for buffering, extraction (micropillars co-flow extraction) and selective adsorption on the MIP core-shell particles.
A miniaturised optical assembly for low-light fluorescence measurements was also developed. Based on small opto-electronic parts and optical fibres, the emission from the MIP particles upon addition of pTyr concentrations from 0.5 – 200 μM could be monitored in real-time.
The reliable identification and quantification of phosphorylated amino acids, peptides and proteins is one of the key challenges in contemporary bioanalytical research, noteworthy, to diagnose and treat diseases at an early developmental stage1. Miniaturised sensing devices like microfluidic chips combined with “smart” detection chemistry, simple data assessment, processing and presentation are very attractive for benchtop use in clinical environments.
We developed novel synthetic probes targeting phosphorylated amino acids, based on core-shell microparticles consisting of a silica core coated with a molecularly imprinted polymer (MIP) shell. These “plastic antibodies” are extremely robust, resist denaturing solvents and elevated temperatures, can be reproducibly produced at low cost, and potentially overcome many of the practical problems in current bioanalytical detection strategies. The MIP layer contains a fluorescent probe monomer, binds selectively to phosphorylated tyrosine (pY) with a significant imprinting factor higher than 3.5 and responds with a “lighting-up” of its fluorescence accompanied by the development of a strongly red-shifted emission band toward the analyte. In analogy to our previous work4, the bead-based ratiometric detection scheme has also been successfully transferred to a microfluidic chip format to demonstrate its applicability to rapid assays. Such a miniaturised device could yield an automated pY measurement system in the future. The setup was built by coupling a modular microfluidic system5 for amino acid functionalisation (Fmoc protection) and, as shown in Figure 1, a multi-layer PDMS/Teflon/glass microfluidic chip6 for buffering, extraction (micropillars co-flow extraction) and selective adsorption on the MIP core-shell particles.
A miniaturised optical assembly for low-light fluorescence measurements was also developed. Based on small opto-electronic parts and optical fibres, the emission from the MIP particles upon addition of pY concentrations from 0.5-200 μM could be monitored in real-time.
Methods for the rapid and sensitive detection of target analytes are gaining importance in medical diagnostics and environmental monitoring, in the security, occupational health and safety as well as food sectors. Among all of the methods employed for rapid tests, lateral flow assays (LFAs) are the most commonly used, and hundreds of test kits based on this technique are available on the market. A major drawback is that most of these capture agents either indicate the analyte only indirectly, and in most cases a second binding agent able to bind directly or indirectly to the analytes is necessary (e.g., a secondary labeled antibody). Furthermore, in certain cases in which the (ultra)trace detection of an analyte is required, the traditional approach of a certain number of probe molecules being conjugated to a particular support is not sufficient. Therefore, novel concepts implementing steps of effective signal amplification are urgently required.
Keeping in mind these limitations, we thought that the sensitivity of these systems should be improvable through employment of gated reporter molecule-releasing hybrid nanoparticle materials on novel lateral flow devices. On one hand, the gated sensor material can produce a massive signal amplification, by releasing many reporter molecules only after chemical recognition of a few analyte molecules has taken place in an independent and separate step at the pore openings. On the other hand, the employment of tailored capture materials for the selective interaction with the released reporter molecules in a second arbitrary zone on the strip allows to concentrate or focus the latter for more efficient detection or to create selective multi-spot detection zones, which renders the simultaneous detection of several reporter molecules at the same time in multiplexed detection of various analytes possible.
For that purpose, we have prepared several stimuli-responsive materials for small-molecule sensing based on specific interactions between biomolecules such as antibodies with the corresponding analytes for the detection of certain explosives. In order to prepare these bio-capped materials, we have selected silica mesoporous nanoparticles (MSNs) as inorganic support due to their unique properties such as defined void structure, high inner surface area and flexible functionalization chemistry. These MSNs are loaded with a brightly fluorescent indicator dye, and the external surface is subsequently functionalized with suitable molecules able to interact with antibodies, efficiently inhibiting dye release. The opening protocol and delivery of the entrapped dye is reminiscent of a displacement reaction involving the presence of the target analyte, producing a displacement of the biomolecule and allowing the detection of the target analyte.
The presentation discusses general aspects of system design as well as analytical performance and highlights the integration into a lateral-flow assay, showing as an example the determination of the explosives TATP, TNT and PETN with fluorescence readout, in single-substance and multiplexing modes.
The tendency of boron-dipyrromethene (BODIPY) dyes to associate in water is well known, and usually a cause for inferior fluorescence properties. Synthetic efforts to chemically improve BODIPYs’ water solubility and minimize this problem have been numerous in the past. However, a deeper understanding of the phenomena responsible for fluorescence quenching is still required. Commonly, the spectroscopic behaviour in aqueous media has been attributed to aggregate or excimer formation, with such works often centring on a single BODIPY family. Herein, we provide an integrating discussion including very diverse types of BODIPY dyes. Our studies revealed that even subtle structural changes can distinctly affect the association behaviour of the fluorophores in water, involving different photophysical processes. The palette of behaviour found ranges from unperturbed emission, to the formation of H or J aggregates and excimers, to the involvement of tightly bound, preformed excimers. These results are a first step to a more generalized understanding of spectroscopic properties vs. structure, facilitating future molecular design of BODIPYs, especially as probes for biological applications.
Gasoline adulteration is a frequent problem world-wide, because of the chance of quick, maximized profits. However, addition of cheaper ethanol or hydrocarbons like kerosene does not only result in economic damage but also poses problems for vehicles and the environment. To enable law enforcement forces, customers or enterprises to uncover such a fraudulent activity directly upon suspicion and without the need to organize for sampling and laboratory analysis, we developed a simple strip-based chemical test. Key to the favorable performance was the dedicated materials tailoring, which led to test strips that consisted of a cellulose support coated with silica, passivated with hexamethyldisilazane and functionalized covalently with a molecular probe. The probe fluoresces brightly across a broad solvent polarity range, enabling reliable quantitative measurements and data analysis with a conventional smartphone. The assays showed high reproducibility and accuracy, allowing not only for the detection of gasoline adulteration but also for the on-site monitoring of the quality of commercial E10 gasoline.
The field of epigenetics describes the relationship between genotype and phenotype, by regulating gene expression without changing the canonical base sequence of DNA. It deals with molecular genomic information that is encoded by a rich repertoire of chemical modifications and molecular interactions. This regulation involves DNA, RNA and proteins that are enzymatically tagged with small molecular groups that alter their physical and chemical properties. It is now clear that epigenetic alterations are involved in development and disease, and thus, are the focus of intensive research. The ability to record epigenetic changes and quantify them in rare medical samples is critical for next generation diagnostics. Optical detection offers the ultimate single-molecule sensitivity and the potential for spectral multiplexing. Here we review recent progress in ultrasensitive optical detection of DNA and histone modifications.
The fluorescence quantum yield (QY) of CdSe dot/CdS rod (DR) nanoparticle ensembles is dependent on the Shell growth and excitation wavelength. We analyze the origin of this dependency by comparing the optical properties of DR ensembles to the results obtained in single-particle experiments. On the Ensemble level, we find that the QY of DRs with shell lengths shorter than 40 nm exhibits no dependence on the excitation wavelength, whereas for DRs with shell lengths longer than 50 nm, the QY significantly decreases for excitation above the CdS band gap. Upon excitation in the CdSe core, the ensemble QY, the fluorescence wavelength, and the fluorescence blinking behavior of individual particles are only dependent on the radial CdS shell thickness and not on the CDs shell length. If the photogenerated excitons can reach the CdSe core region, the fluorescence properties will be dependent only on the surface passivation in close vicinity to the CdSe core. The change in QY upon excitation above the band gap of CdS for longer DRs cannot be explained by nonradiative particles because the ratio of emitting DRs is found to be independent of the DR length. We propose a model after which the decrease in QY for longer CdS shells is due to an increasing fraction of nonradiative exciton recombination within the elongated shell. This is supported by an effective-mass-approximation-based calculation, which suggests an optimum length of DRs of about 40 nm, to combine the benefit of high CdS absorption cross section with a high fluorescence QY.
In all fluorescence-based techniques, the measured signals contain not only sample-related but also instrument-specific contributions, which limit the direct comparison of fluorescence data obtained e.g. on different devices or at different times and often hamper quantification. To rule out instrumentation as major source of variability of emission data, accepted fluorescence standards and procedures for the control of instrument specifications and long-term performance are required. For flow cytometry (FCM), a broad variety of fluorophore-stained polymer beads differing in emission wavelength and intensity is available for the testing of the alignment, sensitivity, and other parameters of FCM. These calibration tools are intended to facilitate the assessment of instrument performance to ensure reliable measurements and to improve the comparability of FCM experiments.
As a step towards an improved comparability of fluorescence data, with special emphasis on spectroscopic methods measuring nano- and micrometer-sized fluorescent objects, we are currently developing a set of fluorescent polystyrene (PS) beads loaded with luminophores from the certified BAM-Kit “Spectral fluorescent standards”, initially developed for the calibration of fluorescence spectrometers. Here, we present first results from studies of these fluorophore-loaded polymer beads. Moreover, new beads are made to supplement this kit by encapsulating near-infrared (NIR)-emissive luminophores in PS beads to cover the UV/VIS, and NIR wavelength range.
These beads are designed for calibration of flow cytometers and other fluorescence imaging systems to meet the increasing demand for reliable and comparable fluorescence data especially in strongly regulated areas like e.g. medical diagnostics.