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Aptamères et électrophorèse capillaire : caractérisation physico-chimique d'aptamères libres en solution ou greffés sur des nanoparticules, et étude de leur affinité avec une cible protéique en vue de leur emploi pour des méthodes sensibles de diagnostic

Abstract : Aptamers are short oligonucleotides selected by SELEX (systematic evolution of ligands by exponential enrichment) and showing great affinity and specificity towards their target. This work deals with the physico-chemical characterization of aptamers and the study of their affinity towards their target, using capillary electrophoresis, through the example of an aptamer selected against a highly basic protein, lysozyme. After having evaluated several modifications of the capillary surface, the permanent coating with hydroxypropylcellulose was chosen in order to minimize protein adsorption while being compatible with the aptamer analysis. The electrophoretic behavior of the aptamer in presence of various cations in solution was then studied using affinity capillary electrophoresis (ACE), showing in case of a dication a significant interaction which could induce a conformational change of the aptamer, and thus influence the interaction with the target. The development of frontal analysis continuous microchip electrophoresis (FACMCE) with fluorescence detection enabled to determine the interaction parameters between lysozyme and its aptamer, as well as the influence of experimental conditions on the binding affinity. Lastly, magnetic fluorescent core-shell nanoparticles were fonctionnalized with the aptamer and characterized by capillary zone electrophoresis (CZE). The FACMCE analysis of the binding between the grafted aptamer and lysozyme showed an affinity similar to those of the free aptamer, enabling to consider the future use of these objects as molecular recognition tools.
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https://pastel.archives-ouvertes.fr/pastel-00553765
Contributor : Marie Girardot <>
Submitted on : Sunday, January 9, 2011 - 4:06:43 PM
Last modification on : Wednesday, October 14, 2020 - 3:43:08 AM
Long-term archiving on: : Saturday, December 3, 2016 - 2:28:53 AM

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  • HAL Id : pastel-00553765, version 1

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Marie Girardot. Aptamères et électrophorèse capillaire : caractérisation physico-chimique d'aptamères libres en solution ou greffés sur des nanoparticules, et étude de leur affinité avec une cible protéique en vue de leur emploi pour des méthodes sensibles de diagnostic. Chimie analytique. Université Pierre et Marie Curie - Paris VI, 2010. Français. ⟨pastel-00553765⟩

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