Celltiter glo 2.0 cell viability assay reagent

The cells were seeded at 2 × 10⁴/well in 48-well cell culture plates. Cell viability was measured using the CellTiter-Glo® 2.0 Cell Viability Assay reagent (Promega, G9243), and the experiment was carried out in accordance with the manufacturer’s protocol. The lactate dehydrogenase (LDH) release assay was performed using a Cytotoxicity Detection Kit (Roche, 11644793001). Calculations of LDH release were performed according to the manufacturer’s protocol.

After the cells were cultured for 24 to 60 h, a volume of 10 µL/well of the Cell Counting Kit-8 assay solution (Dojindo Laboratories, Kumamoto, Japan) was added and the cells were further incubated for 1 h. Absorbance was then measured at 450 nm and 650 nm using a microplate reader. The net difference in absorbance (A450–A650) was used as a measure of metabolic activity. The metabolic activity of the siRNA control-treated cells was considered to be 100%.
After the cells were cultured for 24 to 60 h, an equal amount of the CellTiter-Glo 2.0 Cell Viability Assay reagent (Promega, Madison, WI, USA) was added to each well and incubated for 10 min. Luminescence was measured using a luminometer to determine the amount of ATP, as well as cell proliferation. The amount of ATP in the siRNA control-treated cells was considered to be 100%.