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In the focus of division Biophotonics are the design, preparation, analytical and spectroscopic characterization, and application of molecular and nanoscale
functional materials, particularly materials with a photoluminescence in the visible, near infrared (NIR) and short-wave infrared (SWIR). This includes optical reporters for bioimaging and sensing, security and authentication barcodes, and materials for solid state lighting, energy conversion, and photovoltaics. For the identification of optimum particle structures quantitative spectroscopic studies are performed under application-relevant conditions, focusing on the key performance parameter photoluminescence quantum yield. In addition, simple, cost-efficient, and standardizable strategies for quantifying functional groups on the surface of nano- and microparticles are developed, here with a focus on optical assays and electrochemical titration methods, cross-validated by more advanced methods such as quantitative NMR. In addition, reference materials and reference products are developed for optical methods, particularly luminescence techniques, and for analytical methods utilized for the characterization of nanomaterials.
The development of a Lab-on-a-Chip (LoC) is presented, which can detect reactive phosphorous compounds in the gas phase in combination with an optochemical hand-held sensor. The LoC prototype contains three pairs of sensing materials containing fluorescent indicator dyes in various carrier materials. By measuring the fluorescence response to phosphoryl chloride, a surrogate compound, the detection of chemical warfare agents (CWAs) in gas phase becomes possible within seconds, introducing a novel approach to CWA detection.
PFAS Sensors
(2023)
This contribution provides an introduction to the development of sensors for PFAS analysis, presents the most common approaches, and describes the opto-microfluidic strategy in combination with polymerizable indicators and detection matrices currently being pursued by the Chemical and Optical Sensing Division at BAM.
Per- and polyfluoroalkyl substances (PFAS) are a class of man-made organo-fluorine chemicals that have become environmental contaminants of emerging concern, originating from a variety of materials such as adhesive, stain- and oil-resistant coatings, firefighting foams, etc. The high strength of this C-F bond makes PFAS thermodynamically stable and resistant to (bio)degradation, thus retaining them in the environment over time. Perfluoroalkyl carboxylic acids (PFCAs), one category of the most used PFAS, consist of a fully fluorinated carbon backbone and a charged carboxylic acid headgroup, and have been classified as Substances of Very High Concern (SVHC) and added to the REACH Candidate List due to their persistence in the environment, non-biodegradability and toxicological effects.[1-2] Traditional techniques for the analysis of PFCAs include GC-MS, HRMS and HPLC-based approaches, which are laborious, not portable, costly and require trained personnel. In contrast, fluorescence assays can be designed as easy-to-operate, portable and cost-effective methods with high sensitivity and fast response. Integration of fluorescent probes with an adequately miniaturized assay enables a promising alternative for PFCAs analysis.
Here, a novel guanidine fluorescent probe has been synthesized and fully characterized for the detection of PFCAs in a biphasic extract-&-detect assay. The fluorescent probe was then incorporated into polymeric matrices supported by a red dye-doped SiO2 nanoparticle to construct a dual-emission sensing platform. Such a system allows precise and selective detection of PFCAs, reducing the interference of competitors, matrix effects and other factors except for the PFCAs. The system was then employed in a droplet-based microfluidic setup which offers a portable and easy to operate detection platform.
Die Bundesanstalt für Materialforschung und -prüfung (BAM) ist eine forschende Bundesoberbehörde und Einrichtung der Ressortforschung der Bundesrepublik Deutschland. Unter ihrer Leitlinie „Sicherheit in Technik und Chemie“ ist sie zuständig für die öffentliche technische Sicherheit und für metrologische Aufgaben in der Chemie. Das Aufgabenspektrum der BAM, das sich an aktuellen Fragestellungen aus Wissenschaft, Wirtschaft, Politik und Normung orientiert, bietet sehr viele interessante Tätigkeitsfelder für Naturwissenschaftler*Innen und Ingenieur*Innen.
In recent years, the demand for reliable, versatile, fluorescent pH and oxygen sensors has increased rapidly in many biomedical applications since these analytes are important indicators of cell function or certain diseases. Therefore, sensor particles are needed that are small enough to penetrate cells, non-toxic, and allow for close-up optical monitoring. When developing such sensor systems, one must consider the pH and oxygen range detectable by the sensor dye and the matrix material of the used carrier particles. Here, we present the development of pH- and oxygen-responsive polymeric beads functionalized with fluorescent dyad molecules that consist of an analyte-responsive fluorophore and an analyte-inert dye.
Fluorescent semiconductor nanocrystals, also known as quantum dots (QDs), enabled many advancements in biotechnology, photovoltaics, photocatalysis, quantum computing and display devices. The high versatility of this nanomaterial is based on their unique size-tunable photoluminescence properties, which can be adjusted from the visible to the near-infrared range. In contrast to other nanomaterials, QDs made the transition from a laboratory curiosity to the utilization in commercial products, like the QLED television screen or in smartphone displays. The best investigated QDs are composed of heavy metals like cadmium or lead, which is not the best choice in terms of toxicity and environmental pollution. A more promising material is Indium Phosphide (InP), which is also currently used by Samsung, Sony and co. in the QLED displays.
In this contribution, I would like to give you a sneak peek behind the curtains of nanomaterial synthesis and show how this material is produced, how to stabilize their structural properties, and assess their toxicity in environmentally relevant conditions. Furthermore, I would like to present a synthesis method to accomplish the last open challenge in display technology of a blue luminescent LED based on QDs by introducing a new element to the InP QDs.
The overall interest in nanotoxicity, triggered by the increasing use of nanomaterials in the material and life sciences, and the synthesis of an ever increasing number of new functional nanoparticles calls not only for standardized test procedures1,2 and for efficient approaches to screen the potential genotoxicity of these materials, but also for standardized and validated methods for surface analysis.
4,5 The analysis and quantification of surface chemistry is hence in the focus of an increasing number of standardization organizations and interlaboratory comparisons with different analytical methods are being done.5 For the monitoring of nanomaterial synthesis and the fast assessment of the number of functional groups such as carboxyl and amino functionalities, which are very commonly used functionalities in the life sciences, simple and validated methods are needed that can be performed with common laboratory instrumentation. 5,6 Here we provide a brief overview of the ongoing research in division Biophotonics employing quantitative NMR (qNMR), conductometry, and colorimetric and fluorometric optical assays for the determination of the total and the accessible number of carboxyl and amino groups on differently sized polymer and silica nano- and microparticles.5-7
The reversible analyte-induced switching between a colourless leuco form of a dye and its coloured all-π-conjugated form is one of the oldest concepts in probe- or indicator-based optical analysis,1 constituting so-called “one-color indicators”.2 In contrast to colour changes in the visible region, for which usually a bond-forming or a non-covalent interaction between a functional group on an indicator and an analyte is responsible, the extraordinarily large shifts of 150–250 nm seen for leuco-to-all-π-conjugated transformation are only possible when the reaction takes place directly at a site that is an intrinsic part of a dye’s π-system; the classic case are triphenylmethane dyes.3 Despite its obvious potential, this approach has not been a very popular area of scientific research for decades, perhaps because it is much more difficult to implement selectivity than by modifying terminal functional groups. However, in recent years, reaction-based fluorescence probes have attracted increasing attention by virtue of their superior sensitivity.4,5
Nowadays, rhodamines and fluoresceins are by far the most prominent groups of reaction-based indicators.5 They are usually switched between a colourless and non-fluorescent and a greenish-yellowish absorbing and yellowish-reddish emitting form. Typically, the coloured and fluorescent form is ionic, while the colourless form is neutral, reactions having mainly be designed to occur at the end groups of the xanthenoid π-system. BODIPY dyes, with their favourable spectroscopic and chemical properties as well as facile wavelength tunability features have only very recently been explored into this direction.6,7 Reaction at the core dipyrrin framework of these dyes led to strong colour and fluorescence modulations with potential applications in materials sciences6 and cell imaging.7 The present contribution will highlight the mechanisms at play and the sensing performance realized so far, and will compare core-reactive approaches to reaction-based signalling that involves π-extension of BODIPYs.8,9
The microbial contamination of fuels by fungi or bacteria presents risks such as corrosion and fuel system fouling, which can produce critical problems in refineries and distribution systems and has a significant economic impact at every phase of the process. Many factors have been cited as responsible for microbial growth, like the presence of water in the storage tanks. Indeed, only 1 % water in a storage system is enough for the growth of microorganisms like bacteria or yeasts, as well as for the development of fungal biomass at the oil/water interface.1
In this work, a rapid test for the accurate determination of genomic DNA from aqueous fuel extracts is presented. The detection is based on the employment of polystyrene-mesoporous silica core-shell particles, on which modified fluorescent molecular beacons are covalently grafted. Those beacons incorporate in the hairpin loop a target sequence highly conserved in all bacteria, corresponding to a fragment of the 16S ribosomal RNA subunit. The designed single-stranded molecular beacon contained fluorescein as an internal indicator and a quencher in its proximity when not hybridized. Upon hybridization in presence of the target sequence, the indicator and the quencher are spatially separated, resulting in fluorescence enhancement. To perform the assay the designed particles were disposed on different glass fiber strips to try to achieve a portable and sensitive rapid test. Assays showed that presence of genomic DNA extracts from bacteria down to 50–70 μg L-1 induces a fluorescence response. The optical read-out was adapted for on-site monitoring by adapting a 3D-printed case on a conventional smartphone, taking advantages of the CMOS detector sensitivity.2 Such embedded assembly allowed to detect genomic DNA in aqueous extracts down to the mg L-1 range and presents an interesting step toward on-site monitoring of fuel contamination.
Introduction. Comparing different emitter classes and rationally designing the next generation of molecular and nanoscale probes for bioimaging applications require accurate and quantitative methods for the measurement of the key parameter photoluminescence quantum yield f.1 f equals the number of emitted per number of absorbed photons. This is particularly relevant for increasingly used fluorescence imaging in the short wave-infrared region (SWIR) ≥ 900 nm providing deeper penetration depths, a better image resolution, and an improved signal-to-noise or tumor-to-background ratio.2, 3 However, spectroscopic measurements in the SWIR are more challenging and require specific calibrations and standards.
Lanthanide-based upconversion nanoparticles (UCNPs) like hexagonal 𝛽-NaYF4 UCNPs doped with Yb3+ and Er3+, which efficiently convert 976 nm light to ultraviolet, visible, and near infrared photons, offer new strategies for luminescence-based sensing, barcoding, and Imaging. Their upconversion (UC) luminescence (UCL) features like UCL intensity, quantum yield, relative spectral distribution / UCL luminescence color, and luminescence decay kinetics are, however, strongly influenced by particle size, dopant ion concentration, particle architecture, surface chemistry including presence and thickness of surface passivation and shielding shells, microenvironment/presence of quenchers with high energy vibrations, and excitation power density (P).
We present here a comprehensive study of the influence of excitation power density on the UCL features of different types of UCNPs, focusing on Yb3+ and Er3+ co-doped NaYF4 core-only and core-shell nanostructures with different sizes and doping ion concentration, which underlines the importance of P-dependent optimum dopant concentrations for UCNP performance and the potential of P-tuning of UCL.
A brief perspective of BAM on nanocarriers is presented including examples with special emphasis on the characterization of such materials and underlying challenges. In this respect, also ongoing activities at BAM on different types of core/shell nanomaterials and related systems are briefly summarized.
The use of inorganic lanthanide-doped upconversion nanoparticles (UCNP) in bioimaging and cellular studies requires biocompatible particles. One possible cause of UCNP toxicity is the release of potentially harmful fluoride and lanthanide ions as revealed by dilution studies in aqueous environments, particularly under high dilution conditions. To address this issue, suitable surface coatings preventing such effects in combination with fast screening methods suited for online monitoring and in situ analyses are desired.
Here we present systematic studies of differently sized β-NaYF4:Yb,Er UCNP stabilized with different surface coatings and hydrophilic ligands varying in binding strength to the particle surface in various aqueous environments at different temperatures and UCNP concentrations. The concentration of the fluoride and lanthanide ions released upon particle dissolution was quantified electrochemically with a fluoride ion-sensitive electrode and inductively coupled plasma optical emission spectrometry (ICP-OES) and monitored fluorometrically, thereby exploiting the sensitivity of the upconversion luminescence to changes in size and surface chemistry. Moreover, changes in surface chemistry were determined with X-Ray photoelectron spectroscopy (XPS). Based upon our results, we could derive optimum screening parameters for UCNP stability studies and determine conditions and coating procedures and ligands for enhancing UCNP stability in aqueous environments.
A brief introduction to fluorescence spectroscopy will be provided, ranging from typically measured fluorescence quantities over instrument-specific contributions to measured fluorescence signals to selected applications. In this context, an overview of the photoluminescence properties of molecular and nanoscale luminescence reporters will be given including a brief insight into their photophysics and fluorescence standards designed by division Biophotonics for the calibration and instrument performance validation of fluorescence measuring devices will be presented.
A brief introduction to fluorescence spectroscopy will be provided, ranging from typically measured fluorescence quantities over instrument-specific contributions to measured fluorescence signals to selected applications. In this context, an overview of the photoluminescence properties of molecular and nanoscale luminescence reporters will be given including a brief insight into their photophysics and fluorescence standards designed by division Biophotonics for the calibration and instrument performance validation of fluorescence measuring devices will be presented.
Research of division Biophotonics at the Federal Institute for Materials Research and Testing (BAM) covers several topics including photophysics of molecular and nanocrystalline emitters, the development of signal enhancement, multiplexing, and barcoding strategies, surface group quantification, the rational design of different types of stimuli-responsive optical probes, and concepts and reference materials for the validation of optical-spectroscopic measurements. In the following representative examples for each of these topics are given. Also, current developments like single particle spectroscopy and flow cytometry with lifetime detection and newly certified fluorescence quantum yield standards are presented.
Spectral encoding of cells or particles and the discrimination of multiple spectral codes are a critical process in flow cytometry (FCM). Typical issues in spectral encoding are, e.g., the spectral overlap of codes, or the increasing complexity of instruments . The exploitation of the photoluminescence lifetime (LT) as an encoding parameter could be used to circumvent both of these issues, as it adds another dimension to the parameter space, or, when used as a stand-alone parameter, requiring only one excitation light source and one detector. While LT encoding was considered already decades ago it is still not implemented as a routine technique in FCM yet, mainly due to the challenge of very few photons being available within the limited transition time of a cell or particle through the laser spot.
Recently, we demonstrated LT-FCM based on luminophores with ns LTs in a compact and low-cost flow cytometer. Measurements on polymer microbeads containing luminophores with distinctly different excited state LTs enabled the complete discrimination of three LT codes and five codes in total could be identified.
Now, we have extended our approach towards considerably longer LTs by custom-made polymer microbeads loaded with different ratios of InP/ZnS and AgInS2 quantum dots. The use of these materials significantly expands the usable time range for LT encoding to up to several hundred ns. Our studies demonstrate the possibility to further increase the number of viable LT codes for multiplexing in LT-FCM without the need for extensive hardware modifications.