The BMSCs in the logarithmic growth phase were taken, uniformly inoculated into a 96-well plate at 1x104 cells/well, and added with MTA diluted into different concentrations, with 6 replicates set for each concentration gradient. The cells were further cultured in an incubator for 72 h, followed by the discarding of the original medium. Next, the cells were added with 20 µl of CCK-8 reaction solution (Dojindo) and 170 µl of cell medium, incubated in a dark place at 37˚C for 2 h, and shaken on a micro-vibrator for 3 min. The absorbance at a wavelength of 450 nm was measured using a microplate reader (Detie, Nanjing Detie Laboratory Equipment Co., Ltd.).
Cck 8 reaction solution
Manufactured by Dojindo Laboratories
Sourced in Japan
CCK-8 reaction solution is a cell counting kit used to measure cell viability and proliferation. It is a colorimetric assay based on the reduction of tetrazolium salt WST-8 to a water-soluble formazan dye by dehydrogenase enzymes in living cells. The amount of formazan dye generated is directly proportional to the number of living cells.
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10 protocols using cck 8 reaction solution
1
Cell Viability Assay Using MTA
2
Cell Proliferation Assay Protocol
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The cells were collected and inoculated into 96-well plates at 3 × 103 cells/well. After adhered to the wall, CCK-8 reaction solution (341–07761, Dojindo Laboratories, Kumamoto, Japan) was added at 24, 48, and 72 h, and then incubated for 2 h. The optical value value was measured at 450 nm using a microplate reader. Seven parallel wells were set.
3
Cell Viability Assay of Pancreatic Cancer Cells
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To analyze cell viability, BxPC-3 and PANC-1 cells were seeded at a concentration of 8×103 per 96-well. On the next day, cells were supplied with a fresh medium containing Baicalein, inhibitors, or DMSO (control) or transfected with siRNAs. After 24 h, 48 h, and 72 h of cultivation, 10 μl of the CCK8 reaction solution (Dojindo Laboratories, Japan) was added to each well. After 2-3 h incubation at 37°C in the 5% CO2 incubator, the absorbance at 450 nm was measured by a microplate reader (Sunrise™, Tecan, Männedorf, Switzerland). Moreover, BxPC-3 and PANC-1 cells were transfected with the full-length NEDD9 expression vector. After treated with 50 μM Baicalein for 48 h, cell numbers were counted using an automated cell counter (LUNA™ Automated Cell Counter, Logos Biosystems, Korea).
4
Clonogenic and Viability Assays for Cancer Cells
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500 MDA-MB-231 and MDA-MB-468 cells were seeded into 6-well plates, then routinely cultured in RPMI-1640 medium for 2 weeks. Cells were washed by PBS twice and fixed by methanol, the number of clones was observed under a light microscope. For testing cell viability, 1 × 104 cells were seeded into 96-well plates and cultured for 72 h. 10µL CCK-8 reaction solution (Dojindo, Kumamoto, Japan) was added every 24 h and incubated at 37 °C for 1 h. The absorbance at 450nm was recorded by a microplate reader.
5
Cell Viability Evaluation via CCK-8
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To analyze the cell viability, we performed a CCK-8 assay. MIA PaCa-2 or PANC-1 cells were incubated in a 96-well plate at a final density of 5 × 103 cells/well to allow adherence. After incubation for the specified time, 10 μl CCK-8 reaction solution (Dojindo Laboratories, Kumamoto, Japan) was added to each well. The plates were further incubated at 37 °C for 2 h, and the absorbance was finally determined at 490 nm using a microplate reader.
6
Cell Viability Assay: A2780 Cells
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We incubated A2780 cells for 0, 1, 2, 3, 4, and 5 d, when the CCK-8 reaction solution (Dojindo, Kumamoto, Japan) was added for additional 2 h. The absorbance was recorded to reflect cell viability.
7
Cell Proliferation Assay Protocol
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A2780 and SKOV3 cells were incubated with 3 × 103 cells per well in the 96-well plate. Upon incubation at different time points, 0, 1, 2, and 3 d, each well reacted with the addition of CCK-8 reaction solution (Dojindo, Kumamoto, Japan). Another 2h incubation continued. The optical value was obtained at 450 nm.
8
Cell Viability and Colony Formation Assay
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For cell viability experiment, cells were seeded in a 96-well plate at a density of 2000 cells per well and cultured at 37℃ for 0–7 days. After each time point, 10% CCK-8 reaction solution (DOJINDO, Japan) was added to each well and incubated for another 2 h at 37 °C medium to culture the cells to be measured for 2 h. The absorbance was quantified on a spectrophotometer microplate reader (Multiskan MK3, Thermo) with 450 nm wavelength. Eight experiments were independently conducted every day.
For the colony formation experiment, cells were cultured in 60-mm dish (Corning) with 2000 cells per dish for 14 days. Cells were then fixed with 4% paraformaldehyde (PFA), stained with crystal violet, and analyzed by microscopy.
For the colony formation experiment, cells were cultured in 60-mm dish (Corning) with 2000 cells per dish for 14 days. Cells were then fixed with 4% paraformaldehyde (PFA), stained with crystal violet, and analyzed by microscopy.
9
Cell Proliferation Assay with CCK-8
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The PC cell lines PANC1 and SW1990 were cultured in the 96-well plates at a density of 5,000 cells per well, and 10 μl CCK-8 reaction solution (Dojindo, Japan) was added to each well for 2 h in the incubator with standard culture conditions. The microplate reader (Thermo Fisher Scientific, USA) was used to examine the optical density (OD) values at the absorbance of 450 nm, which represented the relative cell proliferation abilities of the cells.
10
Cell Viability and Colony Formation Assay
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For cell viability experiment, cells were seeded in a 96-well plate at a density of 2000 cells per well and cultured at 37℃ for 0-7 days. After each time point, 10% CCK-8 reaction solution (DOJINDO, Japan) was added to each well and incubated for another 2 h at 37°C medium to culture the cells to be measured for 2 hours. The absorbance was quanti ed on a spectrophotometer microplate reader (Multiskan MK3, Thermo) with 450 nm wavelength. Eight experiments were independently conducted every day.
For the colony formation experiment, cells were cultured in 60-mm dish (Corning) with 2000 cells per dish for 14 days. Cells were then xed with 4% paraformaldehyde (PFA), stained with crystal violet, and analyzed by microscopy.
For the colony formation experiment, cells were cultured in 60-mm dish (Corning) with 2000 cells per dish for 14 days. Cells were then xed with 4% paraformaldehyde (PFA), stained with crystal violet, and analyzed by microscopy.
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