Cell viability was assessed by MTT reduction assay. Briefly, confluent monolayer Vero cells in 96-well plates (15000 cells/well) were incubated for 72 h with different concentrations of the compounds ranging from 0.39 µM to 100 µM in DMEM-1% FCS at 36.8 °C in 5% CO2 atmosphere. Next, cells were incubated for 1 h with 1.0 mg/mL of 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT, PanEco, Moscow, Russia) at 36.8 °C, and then the solution was removed, and precipitated formazan was diluted in DMSO (PanEco, Moscow, Russia). The solution was read on a spectrophotometer at 540 nm. For each concentration, the average cell viability was calculated from the data of three experiments with three replicates each and was expressed as a percentage compared to the untreated cells (100%).
Methyl thiazolyl tetrazolium (mtt)
Manufactured by PanEco
Sourced in Russian Federation
The MTT is a colorimetric assay used for measuring the activity of enzymes that reduce metabolic substrates, indicating cellular viability and proliferation. It is a widely used tool in cell biology research and drug discovery applications.
Automatically generated - may contain errors
Lab products found in correlation
Dimethyl sulfoxide (dmso), by PanEco (3 mentions)
Triton x 100, by Merck Group (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Amphotericin b, by PanEco (1 mentions)
Fetal bovine serum (fbs), by PanEco (1 mentions)
Sodium pyruvate, by PanEco (1 mentions)
Non essential amino acids (neaa), by PanEco (1 mentions)
Trypsin solution, by PanEco (1 mentions)
Penicillin, by PanEco (1 mentions)
Streptomycin, by PanEco (1 mentions)
L glutamine, by PanEco (1 mentions)
Dulbecco modified eagle medium (dmem), by PanEco (1 mentions)
Multiscan fc, by Thermo Fisher Scientific (1 mentions)
6 protocols using methyl thiazolyl tetrazolium (mtt)
1
MTT Assay for Cell Viability
2
MTT Assay for Cell Viability Assessment
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MTT (Thiazolyl Blue Tetrazolium Bromide) was from PanEco (Q105, Russia). MTT was dissolved in PBS to a final concentration 5 mg/mL and filtered through 0.22-μm filter. 10 μL of MTT solution was added to 100 μL of cell culture medium and incubated for 3 h at 37°C, 5% of CO2. After incubation, cell plates were centrifuged at 1500 rpm for 5 min. Media were carefully removed, and 100 μL of pure DMSO was added into each well. The absorbance of the resulting solution was measured at 570 nm by a multi-plate reader.
3
Cadaver Eye Cell Culture Protocol
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Experiments on human cadaver eye tissues were performed in compliance with officially accepted procedures, specifically the Russian Federation law N 4180-I dated 22 December 1992, “On human organs or tissue transplantation” (with the most recent modifications and additions dated 8 December 2020). Human cadaver eyes without ophthalmologic disease were obtained from the Eye Tissue Bank of the S.N. Fyodorov Eye Microsurgery Complex (Moscow, Russia) within 10 h of donor death. Donor age ranged from 30 to 75 years. Collection was conducted in accordance with local ethics requirements, as described in detail previously [25 (link)].
The cells were cultured using reagents from Thermo Fisher Scientific (Thermo Fisher Scientific, Waltham, MA, USA) and culture plastic from Corning (Corning, NY, USA). MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) was produced by the company “PanEco” (PanEco, Moscow, Russian Federation). All reagents for high-performance liquid chromatography (HPLC) were purchased from Sigma-Aldrich (St. Louis, MO, USA) and Fluka (Buchs, Switzerland).
The cells were cultured using reagents from Thermo Fisher Scientific (Thermo Fisher Scientific, Waltham, MA, USA) and culture plastic from Corning (Corning, NY, USA). MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) was produced by the company “PanEco” (PanEco, Moscow, Russian Federation). All reagents for high-performance liquid chromatography (HPLC) were purchased from Sigma-Aldrich (St. Louis, MO, USA) and Fluka (Buchs, Switzerland).
4
Formulation and Evaluation of Lipid-based Nanocarriers
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1,2-Dioleoylphosphatidylcholine (DOPC), 1,2-dioleoylphosphatidylethanolamine (DOPE), and DOTAP chloride (USP grade) were obtained from Lipoid GmbH (Heidelberg, Germany). Sorbitan monostearate (SPAN) was obtained from Sigma-Aldrich (Merk, Buchs, Switzerland), and polysorbate 80 (PS80) and squalene (Sq) were obtained from Acros Organics, and coconut oil triacylglycerols (COTs) were obtained from commercial coconut oil.
DMEM, L-glutamine, sodium pyruvate, non-essential amino acids, penicillin, streptomycin, amphotericin B, MTT, trypsin solution in EDTA, Earle’s solution, glucose, and Versene’s solution were purchased from PanEco, Moscow, Russia. DMSO and Triton X-100 were purchased from Sigma-Aldrich, St. Louis, MO, USA.
Fetal bovine serum was purchased from PanEco, Moscow, Russia.
DMEM, L-glutamine, sodium pyruvate, non-essential amino acids, penicillin, streptomycin, amphotericin B, MTT, trypsin solution in EDTA, Earle’s solution, glucose, and Versene’s solution were purchased from PanEco, Moscow, Russia. DMSO and Triton X-100 were purchased from Sigma-Aldrich, St. Louis, MO, USA.
Fetal bovine serum was purchased from PanEco, Moscow, Russia.
5
MTT Assay for Cell Viability
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Cells were incubated with 0.5 mg/mL MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, PanEco, Moscow, Russia) for 2 h at 37 °C. After that, formazan crystals were extracted from the cells using DMSO (PanEco, Moscow, Russia), followed by shaking for 20 min. The optical density was measured on an xMark plate spectrophotometer (Bio-Rad, Hercules, CA, USA) at a wavelength of 570 nm, subtracting the background value at 620 nm. The results were compared to the control, which was taken as 100%.
6
MTT Assay for Cell Viability
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For this test, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT, Paneco, Moscow, Russia) was prepared as a stock solution of 5 mg/mL in phosphate-buffered saline (PBS, cell culture grade, pH 7.2, Paneco, Moscow, Russia) and filtered. hADMSCs were plated in 96-well plates at a density of 1 × 105 cells per sample. At the days 1, 3 and 5, a solution of 20 µL of MTT was added to each well. After incubation for 4 h at 37 °C, the medium solution was discarded and 100 µL dimethylsulfoxide (DMSO, Paneco, Moscow, Russia) was added to each well, and the absorption was measured by a plate reader Multiscan FC (Thermo Fisher Scientific Inc., Waltham, MA, USA) at 570 nm to determine the cell viability. In this test, viable cells reduce MTT to the purple-colored formazan compound, whereas this reaction will not take place in the dead cells [49 (link)].
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