title: Dimethylformamide interferes with Coomassie dye staining of proteins on blue native gel electrophoresis.
abstract: Blue native gel electrophoresis (BN-PAGE) is used extensively for characterization of mitochondrial respiratory complexes and uses the binding of Coomassie brilliant blue G-250 to visualize proteins. Oxidative modification of sulfhydryl groups of such proteins can be evaluated by labeling with iodoacetamide conjugated to biotin (BIAM) and detected with streptavidin peroxidase on Western blots following BN-PAGE. However, dissolving BIAM in dimethylformamide, a recommended solvent, reduces Coomassie blue G staining to proteins during BN-PAGE. This interference is prevented by dissolving BIAM in dimethyl sulfoxide. Precautions in the use of the dye for protein staining subsequent to BIAM labeling are discussed.
authors.affiliation: Wellcome Trust Research Laboratory, Division of Gastrointestinal Sciences, Christian Medical College, Vellore 632004, India.
authors.forename: V
authors.initials: V
authors.lastname: Raghupathy
authors.affiliation: Neurochemistry Laboratory, Department of Neurological Sciences, Christian Medical College, Vellore 632004, India.
authors.forename: Anna
authors.initials: A
authors.lastname: Oommen
authors.affiliation: Wellcome Trust Research Laboratory, Division of Gastrointestinal Sciences, Christian Medical College, Vellore 632004, India. Electronic address: anup@cmcvellore.ac.in.
authors.forename: Anup
authors.initials: A
authors.lastname: Ramachandran
mesh_terms: D000818:Animals; D001710:Biotin / Q000737:chemistry; D004126:Dimethylformamide / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D007460:Iodoacetamide / Q000737:chemistry; D008931:Mitochondria, Muscle / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000032:analysis; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D051381:Rats; D012394:Rosaniline Dyes / Q000737:chemistry / Q000378:metabolism*; D012997:Solvents / Q000737:chemistry; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D024101:Mitochondrial Proteins; D011506:Proteins; D012394:Rosaniline Dyes; D012997:Solvents; D001710:Biotin; D004126:Dimethylformamide; C004692:coomassie Brilliant Blue; D007460:Iodoacetamide
keywords: Biotin labeling; Blue native electrophoresis; Coomassie blue G