Colorimetric cell proliferation assay for microorganisms in microtiter plate using water-soluble tetrazolium salts.
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.
Tsukatani
Tadayuki
T
Biotechnology and Food Research Institute, Fukuoka Industrial Technology Center, Kurume 839-0861, Japan. tukatani@fitc.pref.fukuoka.jp
Suenaga
Hikaru
H
Higuchi
Tomoko
T
Akao
Tetsuyuki
T
Ishiyama
Munetaka
M
Ezoe
Kimitoshi
K
Matsumoto
Kiyoshi
K
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
D003470:Culture Media; D013778:Tetrazolium Salts