Article (Scientific journals)
Separation and determination of alpha-synuclein monomeric and oligomeric species using two electrophoretic approaches
Napp, Aurore; Houbart, V.; Demelenne, Alice et al.
2018In Electrophoresis, 39 (23), p. 3022-3031
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Keywords :
CGE; Labeling; SDS–PAGE
Abstract :
[en] Parkinson's disease (PD) is a frequent degenerative disorder that is diagnosed based on clinical symptoms. When the first symptoms appear, more than 70% of the dopaminergic cells are already lost. Therefore, it is of utmost importance to have reliable biomarkers to diagnose much earlier PD. In this context, alpha-synuclein (aSyn) is a protein of high interest because of its tendency to form oligomers and amyloid fibrils. The oligomeric forms seem to play a critical pathological role in PD. To date, most of studies aiming at detecting and quantifying aSyn oligomers were performed by immunoassays, mainly by ELISA using specific antibodies. In this study a capillary gel electrophoresis (CGE) coupled with fluorescence detection method was developed to detect and quantify the oligomeric forms of aSyn formed in vitro. All the results obtained were supported by SDS–PAGE analysis, a widely used and well-known technique but exhibiting a main drawback since it is not an automated technique. The repeatability and the intermediate precision of the method were evaluated, as well as the stability of the labeled and non-labeled aSyn samples. After careful screening and optimization of various labeling reagents, 4-fluoro-7-nitrobenzofurazan (NBD-F) was selected and used to establish a calibration curve with monomeric fluorescently-labeled aSyn. Finally, the method was used to study the effect of doxycycline on the oligomerization process. Altogether, our results show that CGE is a very promising automated technique to analyze aSyn monomers, as well as small oligomers. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Napp, Aurore ;  Université de Liège - ULiège > Département de pharmacie > Analyse des médicaments
Houbart, V.;  Laboratory for the Analysis of Medicines, Department of Pharmacy, CIRM, University of LiègeLiège, Belgium
Demelenne, Alice ;  Université de Liège - ULiège > Département de pharmacie > Analyse des médicaments
Merville, Marie-Paule ;  Université de Liège - ULiège > Département de pharmacie > Chimie médicale
Crommen, Jacques ;  Université de Liège - ULiège > Département de pharmacie > Département de pharmacie
Dumoulin, Mireille  ;  Université de Liège - ULiège > Département des sciences de la vie > Département des sciences de la vie
Garraux, Gaëtan  ;  Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Biochimie et physiologie du système nerveux
Servais, Anne-Catherine  ;  Université de Liège - ULiège > Département de pharmacie > Analyse des médicaments
Fillet, Marianne ;  Université de Liège - ULiège > Département de pharmacie > Analyse des médicaments
Language :
English
Title :
Separation and determination of alpha-synuclein monomeric and oligomeric species using two electrophoretic approaches
Publication date :
2018
Journal title :
Electrophoresis
ISSN :
0173-0835
eISSN :
1522-2683
Publisher :
Wiley-VCH Verlag
Volume :
39
Issue :
23
Pages :
3022-3031
Peer reviewed :
Peer Reviewed verified by ORBi
Available on ORBi :
since 07 March 2019

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