title: Antioxidant compounds interfere with the 3.
abstract: Antioxidants are often added to culture media as cytoprotective agents. We examined the effects of antioxidants on the results and interpretation of the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay for cell viability. Without cells, the thiol-containing antioxidant compounds beta-mercaptoethanol, dithiothreitol, pyrrolidine-dithiocarbamate, and N-acetyl-L-cysteine (acetylcysteine) reduced MTT tetrazolium salts to a blue formazan product in a dose-dependent manner. Addition of the compounds L-ascorbic acid and (+)-alpha-tocopherol acid succinate had different effects. In contrast, addition of the antioxidants N-acetyl-5-methoxytryptamine and (-)-2-oxo-4-thiazolidine carboxylic acid, which do not contain reactive thiol groups, did not result in the development of blue formazan product. These results showed that antioxidants, and potentially other chemotherapeutic compounds that contain free thiol groups or other reducing equivalents, readily reduce MTT to produce the blue formazan, irrespective of the viability of the cells present. This undescribed reaction can, therefore, significantly influence the results and interpretation of cell-viability experiments.
authors.affiliation: Department of Radiation Oncology, University of Texas Health Science Center at San Antonio, 78229-3900, USA.
authors.forename: M
authors.initials: M
authors.lastname: Natarajan
authors.forename: S
authors.initials: S
authors.lastname: Mohan
authors.forename: B R
authors.initials: BR
authors.lastname: Martinez
authors.forename: M L
authors.initials: ML
authors.lastname: Meltz
authors.forename: T S
authors.initials: TS
authors.lastname: Herman
mesh_terms: D000111:Acetylcysteine / Q000737:chemistry; D000975:Antioxidants / Q000737:chemistry*; D001205:Ascorbic Acid / Q000737:chemistry; D003124:Colorimetry / Q000379:methods*; D004396:Coloring Agents / Q000737:chemistry*; D003470:Culture Media; D004229:Dithiothreitol / Q000737:chemistry; D007700:Kinetics; D008550:Melatonin / Q000737:chemistry; D008623:Mercaptoethanol / Q000737:chemistry; D010084:Oxidation-Reduction / Q000187:drug effects; D011759:Pyrrolidines / Q000737:chemistry; D011761:Pyrrolidonecarboxylic Acid; D011933:Reagent Kits, Diagnostic; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013778:Tetrazolium Salts / Q000737:chemistry*; D013844:Thiazoles / Q000737:chemistry*; D053778:Thiazolidines; D013859:Thiocarbamates / Q000737:chemistry; D014810:Vitamin E / Q000737:chemistry
chemical_list: D000975:Antioxidants; D004396:Coloring Agents; D003470:Culture Media; D011759:Pyrrolidines; D011933:Reagent Kits, Diagnostic; D013438:Sulfhydryl Compounds; D013778:Tetrazolium Salts; D013844:Thiazoles; D053778:Thiazolidines; D013859:Thiocarbamates; D014810:Vitamin E; C020972:pyrrolidine dithiocarbamic acid; D008623:Mercaptoethanol; C022616:thiazolyl blue; D008550:Melatonin; D001205:Ascorbic Acid; D011761:Pyrrolidonecarboxylic Acid; D004229:Dithiothreitol; D000111:Acetylcysteine; C029536:2-oxothiazolidine-4-carboxylic acid