An improved MTT colorimetric method for rapid viable bacteria counting.

Autor: Xu W; School of Food and Advanced Technology, Massey University, Palmerston North 4410, New Zealand., Shi D; School of Food and Advanced Technology, Massey University, Palmerston North 4410, New Zealand., Chen K; School of Food and Advanced Technology, Massey University, Palmerston North 4410, New Zealand., Palmer J; School of Food and Advanced Technology, Massey University, Palmerston North 4410, New Zealand., Popovich DG; School of Food and Advanced Technology, Massey University, Palmerston North 4410, New Zealand. Electronic address: D.G.Popovich@massey.ac.nz.
Jazyk: angličtina
Zdroj: Journal of microbiological methods [J Microbiol Methods] 2023 Nov; Vol. 214, pp. 106830. Date of Electronic Publication: 2023 Oct 05.
DOI: 10.1016/j.mimet.2023.106830
Abstrakt: The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay has been employed in the analysis of bacterial growth. In comparison to experiments conducted on mammalian cells, the MTT bacterial assay encounters a greater number of interfering factors and obstacles that impact the accuracy of results. In this study, we have elucidated an improved MTT assay protocol and put forth an equation that establishes a correlation between colony-forming units (CFU) and the amount of formazan converted by the bacteria, drawing upon the fundamental principle of the MTT assay. This equation is represented as CFU=kF. Furthermore, we have explicated a methodology to determine the scale factor "k" by employing S. aureus and E. coli as illustrative examples. The findings indicate that S. aureus and E. coli reduce MTT by a cyclic process, from which the optimal reduction time at room temperature was determined to be approximately 30 mins. Furthermore, individual E. coli exhibits an MTT reduction capacity approximately four times greater than that of S. aureus. HPLC analysis proves to be the most accurate method for mitigating interferences during the dissolution and quantification of formazan. Additionally, this study has identified a new constraint related to the narrow linear range (0-125 μg/mL) of formazan concentration-absorbance and has presented strategies to circumvent this limitation.
Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to declare.
(Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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