The MTT colorimetric method [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (Sigma®) involves the absorption of the MTT salt by the cells, which is reduced inside the mitochondria into a product called formazan. This product accumulates inside the cell and is extracted by adding an appropriate solvent [38 (link)]. The cell suspension at a concentration of 3 × 104 cell/mL was distributed in 96-well plates (100 µL per well) and incubated at 37 °C in 5% CO2 for adhesion. Next, 90 µL of culture medium and 10 µL of different treatment concentrations were added, and the cells were incubated again for 24 h. At the end of the incubation period, the MTT solution was added and incubated again for 4 h. The medium was carefully removed, and 100 μL of DMSO was added to solubilize the formazan crystals. The plates were then shaken for 20 min, and the absorbance of each sample was measured at 570 nm using an ELISA reader.
Mtt colorimetric method
Manufactured by Merck Group
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The MTT colorimetric method is a laboratory technique used to assess cell viability and proliferation. It is a quantitative assay that measures the conversion of a tetrazolium salt (MTT) into formazan crystals by metabolically active cells. The intensity of the resulting color change is directly proportional to the number of viable cells, which can be measured spectrophotometrically.
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2 protocols using mtt colorimetric method
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MTT Assay for Cell Viability
2
Mitochondrial Function Assessment by MTT Assay
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To assess the integrity of mitochondrial function, the Tetrazolium 3-(4,5-dimethylthiazol-2-yl)-2,5diphenylbromide (MTT) colorimetric method (SIGMA-ALDRICH, USA) [27] was performed. The analysis takes place by measuring formazan crystal formation. The greater the production of these crystals, the greater the cell viability. After the exposure times, the wells were washed 2 times with 1x PBS (phosphate buffer saline) and 200 µl of the MTT solution at 0.5 mg/ml was added for 3 hours at 37 °C and 5% CO 2 . Then, the reagent solution was removed and 100 µl of the DMSO diluent was added per well, followed by the absorbance reading at 570 nm in a spectrophotometer (Thermo Scientific™ Multiskan™ GO Microplate Spectrophotometer). The percentage of cytotoxicity was obtained by comparing the data with the CTRLgroup according to the equation below:
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