title: Consensus brain-derived protein, extraction protocol for the study of human and murine brain proteome using both 2D-DIGE and mini 2DE immunoblotting.
abstract: Two-dimensional gel electrophoresis (2DE) is a powerful tool to uncover proteome modifications potentially related to different physiological or pathological conditions. Basically, this technique is based on the separation of proteins according to their isoelectric point in a first step, and secondly according to their molecular weights by SDS polyacrylamide gel electrophoresis (SDS-PAGE). In this report an optimized sample preparation protocol for little amount of human post-mortem and mouse brain tissue is described. This method enables to perform both two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) and mini 2DE immunoblotting. The combination of these approaches allows one to not only find new proteins and/or protein modifications in their expression thanks to its compatibility with mass spectrometry detection, but also a new insight into markers validation. Thus, mini-2DE coupled to western blotting permits to identify and validate post-translational modifications, proteins catabolism and provides a qualitative comparison among different conditions and/or treatments. Herein, we provide a method to study components of protein aggregates found in AD and Lewy body dementia such as the amyloid-beta peptide and the alpha-synuclein. Our method can thus be adapted for the analysis of the proteome and insoluble proteins extract from human brain tissue and mice models too. In parallel, it may provide useful information for the study of molecular and cellular pathways involved in neurodegenerative diseases as well as potential novel biomarkers and therapeutic targets.
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Francisco-Jose
authors.initials: FJ
authors.lastname: Fernandez-Gomez
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837; EA 4308-Department of Reproductive Biology-Spermiology-CECOS, CHRU-Lille.
authors.forename: Fanny
authors.initials: F
authors.lastname: Jumeau
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Maxime
authors.initials: M
authors.lastname: Derisbourg
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Burnouf
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Hélène
authors.initials: H
authors.lastname: Tran
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Sabiha
authors.initials: S
authors.lastname: Eddarkaoui
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Hélène
authors.initials: H
authors.lastname: Obriot
authors.affiliation: EA2686-Laboratorie d'Immunologie, Faculté de Médecine - Pôle Recherche.
authors.forename: Virginie
authors.initials: V
authors.lastname: Dutoit-Lefevre
authors.affiliation: Department of Neurology, CHRU-Lille.
authors.forename: Vincent
authors.initials: V
authors.lastname: Deramecourt
authors.affiliation: EA 4308-Department of Reproductive Biology-Spermiology-CECOS, CHRU-Lille.
authors.forename: Valérie
authors.initials: V
authors.lastname: Mitchell
authors.affiliation: EA2686-Laboratorie d'Immunologie, Faculté de Médecine - Pôle Recherche.
authors.forename: Didier
authors.initials: D
authors.lastname: Lefranc
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Malika
authors.initials: M
authors.lastname: Hamdane
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: David
authors.initials: D
authors.lastname: Blum
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Luc
authors.initials: L
authors.lastname: Buée
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837.
authors.forename: Valérie
authors.initials: V
authors.lastname: Buée-Scherrer
authors.affiliation: Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837; nicolas.sergeant@inserm.fr.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Sergeant
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D002232:Carbocyanines / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000378:metabolism; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D057825:Two-Dimensional Difference Gel Electrophoresis / Q000379:methods*
chemical_list: D002232:Carbocyanines; D005456:Fluorescent Dyes; D009419:Nerve Tissue Proteins; D020543:Proteome
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