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Monitoring labeling reactions using fluorescence polarization.
Journal article

Monitoring labeling reactions using fluorescence polarization.

  • Crittin Y Institute of Life Technologies, University of Applied Sciences and Arts Western Switzerland Valais, Route du Rawyl 64, CH-1950 Sion, Switzerland.
  • Héritier J Institute of Life Technologies, University of Applied Sciences and Arts Western Switzerland Valais, Route du Rawyl 64, CH-1950, Switzerland Sion.
  • Segura JM Institute of Life Technologies, University of Applied Sciences and Arts Western Switzerland Valais, Route du Rawyl 64, CH-1950 Sion, Switzerland. jmanuel.segura@hevs.ch.
  • 2014-01-07
Published in:
  • Chimia. - 2013
Anisotropy
Fluorescein-5-isothiocyanate
Fluoresceins
Fluorescent Dyes
Hydrogen-Ion Concentration
Hydrolysis
Molecular Weight
Muramidase
Spectrometry, Fluorescence
Succinimides
Time Factors
Water
English 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.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://sonar.ch/global/documents/273900
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