Wells were prepared and treated in triplicates with increasing concentrations (173 μg/mL, 346 μg/mL, 519 μg/mL and 692 μg/mL) of AELE with one interference well, for 24 h, 48 h or 72 h. For this purpose, AML cells were counted and seeded in 96-well plates at a density of 3 × 105 cells/mL, and were incubated overnight before treatment. The effect of AELE was assayed at these different timeframes using the MTS cell viability reagent (Promega) according to the Manufacturer’s instructions. Cell proliferation was assessed via spectrophotometry by recording the absorbance at a wavelength of 492 nm, using Varioskan™ LUX multimode microplate reader to detect metabolically active cells. Percentage proliferation was calculated by dividing the absorbance of the treated cells with the average absorbance of the control untreated cells. IC50 values were calculated using GraphPad Prism 8.
Mts cell viability reagent
Manufactured by Promega
Sourced in United States
The MTS cell viability reagent is a colorimetric assay used to measure the metabolic activity of cells. It relies on the bioreduction of a tetrazolium compound to produce a colored formazan product that can be quantified spectrophotometrically. The intensity of the color is proportional to the number of viable cells.
Automatically generated - may contain errors
Lab products found in correlation
Plat e cells, by Cell Biolabs (2 mentions)
Cal27, by American Type Culture Collection (1 mentions)
N acetylcysteine nac, by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Ca9 22, by Thermo Fisher Scientific (1 mentions)
Hsc 3, by Thermo Fisher Scientific (1 mentions)
Hsc 3, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Ca9 22, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Prism 8, by GraphPad (1 mentions)
Cisplatin, by Viatris (1 mentions)
5 protocols using mts cell viability reagent
1
Evaluating AELE's Antiproliferative Effects on AML Cells
2
Oral Cancer Cell Line Evaluation
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Oral cancer (CAL 27) cell lines were derived from ATCC (Manassas, VA, USA). Oral cancer (Ca9-22 and HSC-3) cell lines were derived from JCRB Cell Bank (Osaka, Japan). The non-malignant oral cell lines, such as gingival epithelial-derived Smulow–Glickman (S–G) [20 (link),21 ], were used to evaluate the drug safety of SK1. The culture medium for CAL 27, Ca9-22, HSC-3, and S–G cells was a 3:2 mixture of Dulbecco’s Modified Eagle Medium (DMEM) and F12 (Gibco, Grand Island, NY, USA), as previously mentioned [22 (link)].
Cells were treated with SK1 for 24 h. Subsequently, the MTS cell viability reagent (Promega, Madison, WI, USA) was reacted with the cell medium for 1 h to determine cell viability [23 (link)]. To address the function of oxidative stress, N-acetylcysteine (NAC) [24 (link),25 (link),26 (link)] (Sigma-Aldrich, St. Louis, MO, USA) was pretreated (10 mM, 1 h) and SK1 was posttreated for 24 h in different experiments.
Cells were treated with SK1 for 24 h. Subsequently, the MTS cell viability reagent (Promega, Madison, WI, USA) was reacted with the cell medium for 1 h to determine cell viability [23 (link)]. To address the function of oxidative stress, N-acetylcysteine (NAC) [24 (link),25 (link),26 (link)] (Sigma-Aldrich, St. Louis, MO, USA) was pretreated (10 mM, 1 h) and SK1 was posttreated for 24 h in different experiments.
3
Evaluating AELE's Anti-Cancer Potential
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MDA-MB-231 and MCF-7 cells were cultured in 96-well plates in triplicates (density = 1.5 × 105 cells/mL), and were incubated overnight before treatment with, 174, 261, 348, 522, and 696 μg/mL of AELE for 24 h and 48 h. Topotecan (Abcam, 20 μM) [33 (link), 34 (link)] and cisplatin (Mylan, 30 μM) [35 (link), 36 (link)] were used as positive controls. MTS cell viability reagent (Promega) was used to assess the effect of AELE on the cell lines as detailed by Khalil [37 ] . Metabolically active cells were quantified by measuring the absorbance of each well at 492 nm, using Varioskan™ LUX multimode microplate reader. Percentage proliferation was calculated by dividing the absorbance of the treated cells with the average absorbance of the control untreated cells. IC50 values were calculated using GraphPad Prism 8.
4
Immortalized ERCC1 MEF Cell Assay
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Timed matings were performed between Ercc1 heterozygous mice and pregnant females were culled by cervical dislocation at E12.5. Primary MEF cultures were obtained and immortalized using the SV40 large T antigen as described previously23 (link). Briefly, pBABE-SV40-Puro virus was produced using Plat-E cells (Cell Biolabs). 48 hours following transfection, the culture medium containing retroviruses was collected and passed through a 0.22 μm filter. The filtered retrovirus was mixed 1:1 with complete medium and supplemented with 4 μg.ml-1 polybrene, the resulting infective medium was added to primary MEF cultures. Transformed clones were selected for 10 days using 3.5 μg.ml-1 puromycin. Sensitivity to DNA-damaging agents was determined by seeding 500 transformed MEFs per well of a 96-well flat-bottom plate and exposed to mitomycin c (MMC) or ultraviolet (UV) irradiation. After 10 days of culture post-exposure the MTS cell viability reagent (Promega) was added and plates incubated at 37 °C for 4 hours, and absorbance at 492 nm was measured.
5
Immortalized ERCC1 MEF Cell Assay
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