title: Electrophoretic separation of purified myelin: a method to improve the protein pattern resolving.
abstract: Myelin sheath is a lipid-rich membrane, consisting of 70% lipid and 30% proteins, that is involved in physiological and pathological processes. For this reason its protein composition has been often investigated, principally by two-dimensional electrophoresis; however, the consistent lipid content makes it difficult to obtain good proteins separation. To improve the resolution of myelin proteins in a denaturing monodimensional gel electrophoresis, we examined several mixtures for the denaturation of the sample, utilizing different detergents and reducing agents. The definition of the protein pattern was analyzed by both "Blue Silver" Coomassie staining and Western Blot analysis against myelin basic protein, one of the most represented myelin proteins. The best resolution is observed when the sample was incubated with a mixture containing 1.25% dithiothreitol, 4 M urea, and 1% dodecyl maltoside or 1 % 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate, prior to addition of denaturing agents. In conclusion, this work describes a novel method to improve the separation of myelin proteins in a monodimensional gel electrophoresis. It may be also useful for investigating other lipid-rich samples.
authors: Pharmacy Department, University of Genova, Genova, Italy. silvia.ravera@gmail.com
authors: Silvia
authors: S
authors: Ravera
authors: Martina
authors: M
authors: Bartolucci
authors: Giulia
authors: G
authors: Barbarito
authors: Daniela
authors: D
authors: Calzia
authors: Isabella
authors: I
authors: Panfoli
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002793:Cholic Acids / Q000737:chemistry; D003902:Detergents / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005960:Glucosides / Q000737:chemistry; D009185:Myelin Proteins / Q000302:isolation & purification*; D009186:Myelin Sheath / Q000737:chemistry*; D011489:Protein Denaturation; D019163:Reducing Agents / Q000737:chemistry; D014508:Urea / Q000737:chemistry
chemical_list: D002793:Cholic Acids; D003902:Detergents; D005960:Glucosides; D009185:Myelin Proteins; D019163:Reducing Agents; C040358:dodecyl maltoside; D014508:Urea; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate; D004229:Dithiothreitol