Each sample was tested for cell proliferation with the Alamar Blue cell viability reagent assay (Invitrogen, Life Technologies, Grand Valley, N.Y.). Cells collected from each Eppendorf tube were cultured in 48-well plates for 2 weeks. The cell proliferation was determined by supplementing 10% (v/v) Alamar Blue reagent and incubating for another 3 hours before measurement on days 1, 3, 6, 9, 12, and 14. One hundred microliters of supernatant were read at 570/585 nm in a SpectraMax/M2 microplate reader (Molecular Devices, Sunnyvale, Calif.). Cell numbers were determined using the standard curve generated from cultured ADSCs (2, 5, 10, 20, 39, 78, 156, 312, 625, 1,250, 2,500, and 5,000 cells per well).
Alamar blue cell viability reagent assay
Manufactured by Thermo Fisher Scientific
The Alamar Blue cell viability reagent assay is a colorimetric assay that measures the metabolic activity of cells. It utilizes the active ingredient resazurin, which is reduced to the fluorescent compound resorufin by metabolically active cells. The resulting color change or fluorescence can be quantified to assess cell viability, proliferation, and cytotoxicity.
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Lab products found in correlation
Spectramax m2 microplate reader, by Molecular Devices (2 mentions)
Glucose free dmem, by Thermo Fisher Scientific (1 mentions)
Fibronectin, by Merck Group (1 mentions)
Epidermal growth factor (egf), by Merck Group (1 mentions)
Transwell co culture system, by Corning (1 mentions)
Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions)
Transwell plate, by Corning (1 mentions)
3 protocols using alamar blue cell viability reagent assay
1
Alamar Blue Cell Viability Assay
2
Modeling Ischemia-induced Neuronal Injury in vitro
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Oxygen-glucose deprivation (OGD) was used to model an ischemia-like condition in vitro. Cultured cortical neurons were exposed to glucose-free DMEM (Gibco) and the culture plate was placed into a hypoxia incubator (Baker, Sanford, ME) with 0.5% O2, 94.5% N2, and 5% CO2 at 37 °C. After an hour in the OGD, the medium was changed to normal culture neurobasal medium containing B27, 2 mM L-glutamine, and 1% antibiotic solution and plates were returned to normoxia conditions. The cultured cortical neurons following OGD were first treated with NSPCs and/or medium B for 7 days as direct treatment. Transwell co-culture system (8 μm pore size Transwell plates; Corning) was then used for indirect treatment to mimic the in vivo condition. For the Transwell experiment, the cortical neurons following OGD were cultured in the lower chamber and the NSPCs and/or medium B (10 ng/mL EGF (Invitrogen) and 1 µg/mL fibronectin (Sigma)), EGF (10 ng/mL), or fibronectin (1 µg/mL) were added in the upper part of the Transwell inserts. After 7 days, cell viability was assessed using morphological assessment by light microscopy (number of cells per 0.33 mm2) as well as the AlamarBlue Cell Viability Reagent assay (Invitrogen) to analyze the neuronal number.
3
Resveratrol's Effect on Chondrocyte Proliferation
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The effect of resveratrol on chondrocyte proliferation was determined
using alamarBlue cell viability reagent assay (Invitrogen Life Technologies,
Grand Valley, New York). AlamarBlue is a non-toxic aqueous fluorescent
dye that does not affect cell phenotype, viability or cell proliferation.21 (link) Cells were incubated in
96-well plates (5 × 104 cells/well) for four days, to which
resveratrol was added (to make a series of final concentrations:
1, 10, 25 and 50 μmol/l). Each concentration was applied to eight
wells, whereas media supplemented with ethanol served as a control.
After both a 24 hour and 48 hour culture, chondrocytes were supplemented
with 10% (v/v) alamarBlue reagent and incubated for another three
hours. Then, 100 µl of supernatant was read at 570/585 nm in a SpectraMax/M2
microplate reader (Molecular Devices, Sunnyvale, California). Cell
numbers were determined from the standard curve.
using alamarBlue cell viability reagent assay (Invitrogen Life Technologies,
Grand Valley, New York). AlamarBlue is a non-toxic aqueous fluorescent
dye that does not affect cell phenotype, viability or cell proliferation.21 (link) Cells were incubated in
96-well plates (5 × 104 cells/well) for four days, to which
resveratrol was added (to make a series of final concentrations:
1, 10, 25 and 50 μmol/l). Each concentration was applied to eight
wells, whereas media supplemented with ethanol served as a control.
After both a 24 hour and 48 hour culture, chondrocytes were supplemented
with 10% (v/v) alamarBlue reagent and incubated for another three
hours. Then, 100 µl of supernatant was read at 570/585 nm in a SpectraMax/M2
microplate reader (Molecular Devices, Sunnyvale, California). Cell
numbers were determined from the standard curve.
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