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- Fluorescence (3)
- Quantum yield (3)
- Integrating sphere (2)
- Lifetime (2)
- Photoluminescence (2)
- Absolute quantum yield (1)
- Anisotropy (1)
- Dye (1)
- Emission (1)
- Excitation (1)
- Luminescence (1)
- Method (1)
- Method comparison (1)
- Multiparametric (1)
- Multiplexing (1)
- NIR (1)
- NIR dyes (1)
- Nanoparticles (1)
- Quantum dot (1)
- Quantum efficiency (1)
- Quantum yield standards (1)
- Quinine sulfate dihydrate (1)
- Reabsorption (1)
- Rhodamine 101 (1)
- Spectral correction (1)
- Standard (1)
- Standards (1)
- Surface analysis (1)
- Uncertainty (1)
There is an increasing interest in chromophores absorbing and emitting in the near-infrared (NIR) spectral region, e.g., for applications as fluorescent reporters for optical imaging techniques and hence, in reliable methods for the characterization of their signal-relevant properties like the fluorescence quantum yield (Φf) and brightness. The lack of well established Φf standards for the NIR region in conjunction with the need for accurate Φf measurements in transparent and scattering media encouraged us to built up an integrating sphere setup for spectrally resolved measurements of absolute fluorescence traceable to radiometric scales. Here, we present the design of this setup and its characterization and validation including an uncertainty budget for the determination of absolute Φf in the visible and NIR. To provide the basis for better measurements of Φf in the spectral window from ca. 600 to 1000 nm used, e.g., for optical imaging, the absolute Φf of a set of NIR chromophores covering this spectral region are measured and compared to relative values obtained using rhodamine 101 as Φf standard. Additionally, the absolute Φf values of some red dyes that are among the most commonly used labels in the life sciences are presented as well as the absolute quantum yield of an optical probe for tumor imaging.
The commercial availability of stand-alone setups for the determination of absolute photoluminescence quantum yields (φf) in conjunction with the increasing use of integrating sphere accessories for spectrofluorometers is expected to have a considerable influence not only on the characterization of chromophore systems for use in optical and opto-electronic devices, but also on the determination of this key parameter for (bio)analytically relevant dyes and functional luminophores. Despite the huge potential of systems measuring absolute φf values and the renewed interest in dependable data, evaluated protocols for even the most elementary case, the determination of the fluorescence quantum yield of transparent dilute solutions of small organic dyes with integrating sphere methods, are still missing. This encouraged us to evaluate the performance and sources of uncertainty of a simple commercial integrating sphere setup with dilute solutions of two of the best characterized fluorescence quantum yield standards, quinine sulfate dihydrate and rhodamine 101, strongly differing in spectral overlap between absorption and emission. Special attention is dedicated to illustrate common pitfalls of this approach, thereby deriving simple procedures to minimize measurement uncertainties and improve the comparability of data for the broad community of users of fluorescence techniques.
Unter dem Motto „Innovation und Qualitätssicherung in der (Bio)Analytik“ werden in der Arbeitsgruppe Fluoreszenzspektroskopie der BAM, Bundesanstalt für Materialforschung und -prüfung, funktionelle Chromophor-Systeme, einfache Signalverstärkungs- und Multiplexingstrategien sowie innovative Validierungs- und rückführbare Standardisierungskonzepte für verschiedene fluorometrische Messgrößen und Methoden entwickelt. Im Mittelpunkt stehen dabei molekulare Fluorophore, Nanokristalle mit größenabhängigen optischen Eigenschaften (sogenannte Quantenpunkte, QDs) und fluoreszierende Partikel variabler Größe sowie Sonden und Sensormoleküle für neutrale und ionische Analyte und für die Charakterisierung von funktionellen Gruppen. Dabei erfolgen auch methodische Entwicklungen für die Fluoreszenzspektroskopie, die Fluoreszenzmikroskopie, die Milcrofluorometrie, die Sensorik und die Mikroarraytechnologie. Ziele sind u. a. das Design und die Untersuchung von multiplexfähigen selektiven und sensitiven Sonden für die Biomarkeranalytik, die Entwicklung von Methoden zur Charakterisierung der signalrelevanten Eigenschaften dieser Chromophor-Systeme und zur Charakterisierung von funktionellen Gruppen an Oberflächen und ihre Validierung sowie die Entwicklung und Bereitstellung von formatadaptierbaren, flexibel ersetzbaren
Standards für die fluoreszenzbasierte Multiparameteranalytik.