In vitro cell viability was determined with the MTT cell viability assay kit as described by the manufacturer (R&D systems). Briefly, cells were cultured at 15×103 cells per well in 96-well tissue culture plates and incubated at 37°C for 24 or 48 hr with or control IgG (PK136 mAb, 50 µg/mL), 14G2a (50 µg/mL), BQ123 (5 µM), 14G2a (50 µg/mL)+BQ123 (5 µM), or BKM120 (50 µM) for 24 or 48 hr. At the end of the culture period, cells were washed with PBS, the MTT reagents were added according to the manufacturer's recommendations, and the absorbance was measured at 570 nm using an ELISA plate reader. Viability of the control cells was defined as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates.
Mtt cell viability assay kit
Manufactured by R&D Systems
Sourced in United States
The MTT Cell Viability Assay Kit is a colorimetric assay used to measure cell metabolic activity. The assay is based on the conversion of the tetrazolium salt MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) into a formazan product by the mitochondrial enzymes of living cells. The resulting formazan can be quantified by measuring the absorbance at 570 nm, which is proportional to the number of viable cells.
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Lab products found in correlation
Spectramax microplate spectrophotometer, by Molecular Devices (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Promega (1 mentions)
Dexamethasone (dex), by Fujifilm (1 mentions)
Butyric acid, by Fujifilm (1 mentions)
Insulin, by Fujifilm (1 mentions)
3t3 l1 pre adipocytes, by Fujifilm (1 mentions)
Dulbecco modified eagle medium (dmem), by Fujifilm (1 mentions)
Ffa free bovine serum albumin, by Fujifilm (1 mentions)
Dulbecco modified eagle medium (dmem), by Nissui Pharmaceutical (1 mentions)
6 protocols using mtt cell viability assay kit
1
Cell Viability Determination by MTT Assay
2
MTT Cell Viability Assay of Peptides
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Cell viability was determined using the MTT Cell Viability Assay Kit from R&D Systems (Minneapolis, MN, USA) following the manufacturer’s instructions. Briefly, aliquots of 1.5 × 104 Huh7.5/CD81 cells/well were cultured in 96-well plates with fresh medium or medium with increasing concentrations of peptides for 48 h. The absorbance at 490 nm was measured with a 96-well plate reader. Each experiment was repeated at least three times.
3
Evaluating miR-98 Effect on Lung Cancer Proliferation
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Effect of miR-98 on proliferation of lung cancer cells was studied by measuring the relative absorbance of PC9 and A549 cells by using MTT Cell Viability Assay Kit (R&D Systems, Inc., Minneapolis, MN, USA), strictly following the manufacturer’s instructions. Briefly, PC9 and A549 cells were seed in a 96-well tissue culture plate (1×104 cells/mL) and incubated for 4 days. For MTT assay, 15 µL of MTT (5 mg/mL; Promega Corporation, Fitchburg, WI, USA) was added to each well and incubated at 37°C for 4 hours. At the end of the incubation period, the untransformed MTT was removed and 100 µL of dimethyl sulfoxide was added to each well to lyse the cells. The absorbance value of each well was read at a wavelength of 490 nm using a SpectraMax Microplate Spectrophotometer (Molecular Devices Corporation, Sunnyvale, CA, USA). All these assays were performed in triplicate.
4
Macrophage and Adipocyte Cell Culture
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Butyric acid and FFA-free bovine serum albumin (BSA) were purchased from Wako Chemicals (Osaka, Japan). All other chemicals were obtained from Nacalai Tesque (Kyoto, Japan). RAW264.7 mouse macrophages and 3T3-L1 mouse pre-adipocytes (American Type Culture Collection, Manassas, VA) were cultured in DMEM (Nissui Pharmaceutical Co., Ltd., Tokyo, Japan) with 292 μg/mL L-glutamine, 100 units/mL penicillin, 100 μg/mL streptomycin, and 10% fetal bovine serum (FBS) at 37 °C under 5% CO2. Differentiation of 3T3-L1 pre-adipocytes was induced by addition of 0.5 mmol/L 13-isobutyl-1-methylxanthine, 2.5 μmol/L dexamethasone, and 10 μg/mL insulin (Wako Chemical) in DMEM containing 10% FBS, beginning 2 days after the cells reached confluence in a 24-well plate. The medium was then replaced with DMEM containing 10% FBS and 5.0 μg/mL insulin and was changed every 2 or 3 days. Twenty days after induction of differentiation, the hypertrophied 3T3-L1 adipocytes were used in experiments. Cell viability was assessed using an MTT cell viability assay kit (R&D Systems, Minneapolis, MN).
5
Huh7.5.1 Cells Cytotoxicity Assay
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Huh7.5.1 cells were treated with E27 for the indicated concentration and period of time. After removal of E27, cells were incubated for an additional 48 h in 24-well plates. Cytotoxicity was determined using the MTT Cell Viability Assay Kit from R&D Systems (Minneapolis, MN, USA).
6
Cytotoxicity Assay of Compound Treatment
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Huh7.5.1 cells (105 per well) were treated with D1ECL1S+D2ECL2 or 6HV1 for the indicated concentration and period of time. After removal of D1ECL1S+D2ECL2 or 6HV1, cells were incubated for an additional 48 hours in 96-well plates. Cytotoxicity was determined using the MTT Cell Viability Assay Kit from R&D Systems (Minneapolis, MN, USA Cat# 4890-025-K).
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