title: Improved separation of alpha chains of collagen type I, type III, and type V by noninterrupted electrophoresis using thioglycolic acid as a negatively charged reducer.
abstract: Improved separation of alpha chains of collagen type I (alpha 1 [I]2 alpha 2[I]), type III(alpha 1[III]3), and type V (alpha 1[V]alpha 3[V])was achieved by noninterrupted sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a negatively charged reducer, thioglycolic acid. The thioglycolic acid, added to the running buffer of the cathodic reservoir, in the middle of electrophoresis quickly migrated in the gel anode, reducing interchain disulfide linkages in collagen type III and dissociating it into its alpha chain monomer, alpha 1[III], without an interruption of electrophoresis. The alpha chain, alpha 1[III], migrated more slowly than the alpha 1 [I] and alpha 2[I] chains of collagen type I, resulting in an excellent separation of alpha 1[III] from alpha 1[I]. The mobility of alpha 1[III] could be controlled by varying the time of thioglycolic acid addition to the running buffer. This enabled us not only to separate alpha 1[III] from alpha 1[I] and alpha 1[V], but also to precisely quantitate these alpha chains, even at low protein loading of mixed samples.
authors.affiliation: Department of Biochemistry, Department of Neurosurgery, Yamaguchi University School of Medicine, Ube, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Nakamura
authors.forename: S
authors.initials: S
authors.lastname: Inoue
authors.forename: S
authors.initials: S
authors.lastname: Abiko
authors.forename: H
authors.initials: H
authors.lastname: Aoki
authors.forename: K
authors.initials: K
authors.lastname: Takeo
mesh_terms: D003094:Collagen / Q000302:isolation & purification*; D000090602:Collagen Type I, alpha 1 Chain; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006801:Humans; D010084:Oxidation-Reduction; D013864:Thioglycolates
chemical_list: C000716127:COL1A1 protein, human; D000090602:Collagen Type I, alpha 1 Chain; D013864:Thioglycolates; D003094:Collagen