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Self-quenching of uranin: Instrument response function for color sensitive photo-detectors |
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| Authors: | Luchowski Rafal Sabnis Sushant Szabelski Mariusz Sarkar Pabak Raut Sangram Gryczynski Zygmunt Borejdo Julian Bojarski Piotr Gryczynski Ignacy |
| Institution: | aCenter for Commercialization of Fluorescence Technologies (CCFT), Department of Molecular Biology & Immunology, UNTHSC, Fort Worth, TX 76107, USA;bDepartment of Biophysics, Institute of Physics, Maria Curie-Sklodowska University, 20-031 Lublin, Poland;cDepartment of Physics and Biophysics, University of Warmia and Mazury in Olsztyn, 10-719 Olsztyn, Poland;dInstitute of Experimental Physics, University of Gdansk, Wita Stwosza 57, 80-952 Gdansk, Poland;eDepartment of Cell Biology and Anatomy, UNTHSC, Fort Worth, TX 76107, USA |
| Abstract: | Concentration is a key determining factor in the fluorescence properties of organic fluorophores. We studied self-quenching of disodium fluorescein (uranin) fluorescence in polyvinyl alcohol (PVA) thin films. The concentration dependent changes in brightness and anisotropy were followed by a lifetime decrease. We found that at a concentration of 0.54 M, the lifetime decreases to 7 ps. At a concentration of 0.18 M the lifetime was 10 ps with the relatively high quantum yield of 0.002. In these conditions the fluorescence intensity decay was homogeneous (well approximated by a single lifetime). We realized that such a sample was an ideal fluorescence lifetime standard for spectroscopy and microscopy, and therefore characterized instrument response functions for a time-domain technique. We show that self-quenched uranin enables measurements free of the color effect, making it a superior choice for a lifetime reference over scattered light. |
| Keywords: | Self-quenching Homotransfer Homo-FRET Instrument response function Lifetime standard Single photon avalanche diode |
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