title: Colorimetric cell proliferation assay for microorganisms in microtiter plate using water-soluble tetrazolium salts.
abstract: A colorimetric method to assay cell proliferation of microorganisms in 96-well microtiter plates using water-soluble tetrazolium salts and electron mediators was developed. Combinations of 6 kinds of water-soluble tetrazolium salts and 27 kinds of electron mediators that considered the metabolic efficiency of microorganisms and the influence with medium components were investigated. 2-Methyl-1,4-naphthoquinone (NQ) was reduced most effectively by various species of microorganisms, and a combination of WST-8 as a water-soluble tetrazolium salt with 2-methyl-1,4-NQ repressed the increase in background due to medium components. In the presence of 2-methyl-1,4-NQ, WST-8 was reduced by microbial cells to formazan, which exhibited maximum absorbance at 460 nm. The proposed tetrazolium method could be applied to measure proliferations of various microbial cells including 3 kinds of yeast, 9 kinds of Gram-positive bacteria, and 10 kinds of Gram-negative bacteria. Linear relationships between the absorbance and viable microbial cell density were obtained in all microorganisms, suggesting that the absorbance change reflected the microbial cell proliferation.
authors.affiliation: Biotechnology and Food Research Institute, Fukuoka Industrial Technology Center, Kurume 839-0861, Japan. tukatani@fitc.pref.fukuoka.jp
authors.forename: Tadayuki
authors.initials: T
authors.lastname: Tsukatani
authors.forename: Hikaru
authors.initials: H
authors.lastname: Suenaga
authors.forename: Tomoko
authors.initials: T
authors.lastname: Higuchi
authors.forename: Tetsuyuki
authors.initials: T
authors.lastname: Akao
authors.forename: Munetaka
authors.initials: M
authors.lastname: Ishiyama
authors.forename: Kimitoshi
authors.initials: K
authors.lastname: Ezoe
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Matsumoto
mesh_terms: D001419:Bacteria / Q000254:growth & development* / Q000378:metabolism; D049109:Cell Proliferation; D003124:Colorimetry / Q000379:methods; D003470:Culture Media / Q000737:chemistry / Q000378:metabolism; D008828:Microbiological Techniques / Q000379:methods*; D013778:Tetrazolium Salts / Q000737:chemistry / Q000378:metabolism*; D015003:Yeasts / Q000254:growth & development* / Q000378:metabolism
chemical_list: D003470:Culture Media; D013778:Tetrazolium Salts