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The need for the traceable characterization of fluorescence instruments is emphasized from a chemists point of view, focusing on spectral fluorescence standards for the determination of the wavelength- and polarization-dependent relative spectral responsivity and relative spectral irradiance of fluorescence measuring systems, respectively. In a first step, major sources of error of fluorescence measurements and instrument calibration are revealed to underline the importance of this issue and to illustrate advantages and disadvantages of physical and chemical transfer standards for generation of spectral correction curves. Secondly, examples for sets of traceable chemical emission and excitation standards are shown that cover a broad spectral region and simple procedures for the determination of corrected emission spectra with acceptable uncertainties are presented. With proper consideration of the respective measurement principle and geometry, these dye-based characterization procedures can be not only applied to spectrofluorometers but also to other types of fluorescence measuring systems and even to Raman spectrometers.
The inter-instrument, inter-laboratory, and long-term comparability of fluorescence data requires the correction of the measured emission and excitation spectra for the wavelength- and polarization-dependent spectral irradiance of the excitation channel at the sample position and the spectral responsivity of the emission channel employing procedures that guarantee traceability to the respective primary standards. In this respect the traceability chain of fluorometry is discussed from a radiometrists point of view. This involves, in a first step, the realization of the spectral radiance scale, based on the blackbody radiator and electron storage ring, and the spectral responsivity scale, based on the cryogenic radiometer and their control via key comparisons of the national metrology institutes. In a second step, the characterization including state-of-the art uncertainties of the respective source and detector transfer standards such as tungsten strip lamps, integrating sphere radiators, and trap detectors used to disseminate these radiometric quantities to users of spectroscopic techniques is presented.
The development of fluorescence applications in the life and material sciences has proceeded largely without sufficient concern for the measurement uncertainties related to the characterization of fluorescence instruments. In this first part of a two-part series on the state-of-the-art comparability of corrected emission spectra, four National Metrology Institutes active in high-precision steady-state fluorometry performed a first comparison of fluorescence measurement capabilities by evaluating physical transfer standard (PTS)-based and reference material (RM)-based calibration methods. To identify achievable comparability and sources of error in instrument calibration, the emission spectra of three test dyes in the wavelength region from 300 to 770 nm were corrected and compared using both calibration methods. The results, obtained for typical spectrofluorometric (0°/90° transmitting) and colorimetric (45°/0° front-face) measurement geometries, demonstrated a comparability of corrected emission spectra within a relative standard uncertainty of 4.2% for PTS- and 2.4% for RM-based spectral correction when measurements and calibrations were performed under identical conditions. Moreover, the emission spectra of RMs F001 to F005, certified by BAM, Federal Institute for Materials Research and Testing, were confirmed. These RMs were subsequently used for the assessment of the comparability of RM-based corrected emission spectra of field laboratories using common commercial spectrofluorometers and routine measurement conditions in part 2 of this series (subsequent paper in this issue).
The self-assembly of double-hydrophilic poly(ethylene oxide)–poly(2-methyl-2-oxazoline) diblock copolymers in water has been studied. Isothermal titration calorimetry, small-angle X-ray scattering, and analytical ultracentrifugation suggest that only single polymer chains are present in solution. In contrast, light scattering and transmission electron microscopy detect aggregates with radii of ca. 100 nm. Pulsed field gradient NMR spectroscopy confirms the presence of aggregates, although only 2% of the polymer chains undergo aggregation. Water uptake experiments indicate differences in the hydrophilicity of the two blocks, which is believed to be the origin of the unexpected aggregation behavior (in accordance with an earlier study by Ke et al. [Macromolecules2009, 42, 5339–5344]). The data therefore suggest that even in double-hydrophilic block copolymers, differences in hydrophilicity are sufficient to drive polymer aggregation, a phenomenon that has largely been overlooked or ignored so far.
In vielen Umgebungen, besonders bei hohen Temperaturen, korrosiven Umgebungen oder auf bewegten oder schlecht zugänglichen Flächen, kann die Temperatur nicht oder nur mit nicht akzeptablem Aufwand mit Berührungsthermometern gemessen werden. Diese Umgebungsbedingungen sind unter anderem in der chemischen Industrie, der Lebensmittel-, Metall-, Glas-, Kunststoff- und Papierherstellung sowie bei der Lacktrocknung anzutreffen. In diesen Bereichen kommen Strahlungsthermometer zum Einsatz. Der VDI-Statusreport zeigt typische Anwendungsfelder von nicht radiometrisch kalibrierten Wärmebildkameras und von radiometrisch kalibrierten Thermografiekameras. Um verlässlich mit spezifizierten Messunsicherheiten berührungslos Temperaturen zu messen, müssen die Strahlungsthermometer und Thermografiekameras nicht nur kalibriert, sondern radiometrisch und strahlungsthermometrisch umfassend charakterisiert werden. Auch die optische Materialeigenschaft, der spektrale Emissionsgrad und die Gesamtstrahlungsbilanz (Strahlung des Messobjekts und der Umgebung) sind bei der industriellen Temperaturmessung von großer Bedeutung. In den letzten Jahrzehnten ist dazu ein umfassendes technisches Regelwerk entstanden, das wir Ihnen mit diesem VDI-Statusreport vorstellen. Manche in den Richtlinien beschriebenen Kennwerte mögen abstrakt wirken. In diesem Statusreport zeigen wir an konkreten Beispielen, was diese Kenngrößen für die berührungslose Temperaturmessung bedeuten. Beispiele von Anwendungen zeigen, wo temperaturmessende Thermografiekameras und ausschließlich bildgebende Wärmebildkameras in der Praxis eingesetzt werden. Mit einer Analyse, welche Themen und Anwendungen derzeit besonders intensiv diskutiert werden, versuchen wir Trends für zukünftige Entwicklungen herauszuarbeiten.
The phase behavior of a dendritic amphiphile containing a Newkome-type dendron as the hydrophilic moiety and a cholesterol unit as the hydrophobic segment is investigated at the air–liquid interface. The amphiphile forms stable monomolecular films at the air–liquid interface on different subphases. Furthermore, the mineralization of calcium Phosphate beneath the monolayer at different calcium and phosphate concentrations versus mineralization time shows that at low calcium and Phosphate concentrations needles form, whereas flakes and spheres dominate at higher concentrations. Energy-dispersive X-ray spectroscopy, X-ray photoelectron spectroscopy, and electron diffraction confirm the formation of calcium phosphate. High-resolution transmission electron microscopy and electron diffraction confirm the predominant formation of octacalcium phosphate and hydroxyapatite.
The data also indicate that the final products form via a complex multistep reaction, including an association step, where nano-needles aggregate into larger flake-like objects.
In the second part of this two-part series on the state-of-the-art comparability of corrected emission spectra, we have extended this assessment to the broader community of fluorescence spectroscopists by involving 12 field laboratories that were randomly selected on the basis of their fluorescence measuring equipment. These laboratories performed a reference material (RM)-based fluorometer calibration with commercially available spectral fluorescence standards following a standard operating procedure that involved routine measurement conditions and the data evaluation software LINKCORR developed and provided by the Federal Institute for Materials Research and Testing (BAM). This instrument-specific emission correction curve was subsequently used for the determination of the corrected emission spectra of three test dyes, X, QS, and Y, revealing an average accuracy of 6.8% for the corrected emission spectra. This compares well with the relative standard uncertainties of 4.2% for physical standard-based spectral corrections demonstrated in the first part of this study (previous paper in this issue) involving an international group of four expert laboratories. The excellent comparability of the measurements of the field laboratories also demonstrates the effectiveness of RM-based correction procedures.