Monitoring Labeling Reactions Using Fluorescence Polarization
FH-HES Universities of Applied Sciences
DOI:
https://doi.org/10.2533/chimia.2013.740Keywords:
Fluorescence anisotropy, Fluorescence polarization, Fluorescent labeling, LysozymeAbstract
Fluorescence techniques are widely applied in protein research owing to their specificity and sensitivity, but require prior fluorescent labeling. Here we show a novel approach to optimize labeling protocols by monitoring labeling reactions using fluorescence polarization: the larger molecular mass of the fluorescent protein conjugate compared to the dye alone results in an increase in fluorescence anisotropy during the reaction. Thereby, labeling of lysozyme with fluorescein isothiocyanate or carboxyfluorescein succinimidyl ester could be monitored and the influence of parameters such as the pH could be quantitatively assessed. Moreover, the impact and kinetics of side reactions such as hydrolysis were determined. This new method is rapid, easy to implement, and generically applicable.Downloads
Published
2013-10-30
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Copyright (c) 2013 Swiss Chemical Society
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
How to Cite
[1]
Chimia 2013, 67, 740, DOI: 10.2533/chimia.2013.740.