Cell proliferation was measured using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. 48 h after transfection, the cells were seeded at a density of 3×103 cells/well in 96-well plates. At indicated time points, 20 μl MTT regents (5 mg/ml; Sigma-Aldrich, St. Louis, MO, USA) was supplemented into each well, and the plates were incubated for another 4 h. Then the supernatant was removed before the addition of 150 μl DMSO (Sigma-Aldrich). A microplate reader (Bio-Rad Laboratories, Hercules, CA, USA) was used to measure the absorbance at 570 nm.
Mtt regent
Manufactured by Merck Group
Sourced in United States
MTT reagents are a colorimetric assay used to measure cell metabolic activity and cell proliferation. The reagents contain a tetrazolium salt that is reduced by metabolically active cells, producing a colored formazan product that can be measured spectrophotometrically.
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5 protocols using mtt regent
1
MTT Assay for Cell Proliferation
2
Cell Proliferation Assay with MTT
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Cell proliferation ability was evaluated by MTT assay. Briefly, transfected U2OS and MG63 cells were seeded into 96-well plates at a density of 5 × 103/well and routinely cultured for 24 h, 48 h, 72 h and 96 h, respectively. At the indicated time, 20 μL MTT regents (5 mg/mL; Sigma-Aldrich, Irvine, Ayrshire, UK) was supplemented into each well at a final MTT concentration of 0.45 mg⁄mL and incubated for another 4 h at 37 °C in a humidified chamber. Subsequently, the medium was removed and 150 μL of dimethylsulfoxide (DMSO; Sigma-Aldrich) was added to each well to dissolve the blue formazan crystals for 30 min. The absorbance value at 490 nm was recorded on the Model 680 microplate reader (Bio-Rad, Hercules, CA, USA). The measurements for each sample were performed in triplicate.
3
Evaluating H9C2 Cell Viability via MTT Assay
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The viability of H9C2 cells was evaluated via MTT assay. H9C2 cells were planted into 96-well plates. MTT regents (5 mg/ml; Sigma-Aldrich) was supplemented in the culture medium and cells were cultured at 37°C for 4 h in the darkness at 37°C for 4 h. To dissolve the formazan of MTT, dimethyl sulfoxide was then added. The absorbance at 490 nm was read to determine the viability of H9C2 cells.
4
Measuring Cell Proliferation via MTT Assay
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For detection of cell proliferation, MTT assay were performed at 48 h post-transduction. Total 10 μl of MTT regent (Sigma-Aldrich, US) at the concentration of 5 mg/ml was added to each well. Following 3 h of incubation, 100 μl of dimethyl sulphoxide (DMSO) detergent (Sigma-Aldrich, US) was added to each well. The plate was then incubated on a shaker at 4 ℃ for 2 h, and absorbance was measured at a 570 nm with a microplate reader (Bio-Rad, US).
5
Evaluating Cell Viability and Death
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In this study, cell viability was measured through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Annexin V/7-AAD assay [35 (link)]. For MTT assay, cultured cells were first planted into 96-well plates. MTT regent was added at a volume of 10 μL into each well (0.5 mg/mL, Sigma-Aldrich, St. Louis, MO, USA). The plate was incubated at 37 °C for 4 h. Medium was then removed from the plate. Finally, 100 μL DMSO was added into each well to dissolve the crystal. The absorbency at a wavelength of 492 nm was measured by a microplate reader (Allsheng, Hangzhou, China). For cell death assay, Annexin V/7-AAD assay was performed on a flow cytometer (BD Biosciences, USA) to detect the cellular fluorescence using an Annexin V-PE/7-AAD detection kit (Vazyme, Nanjing, China) according to the manufacturers’ protocol.
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