Mtt proliferation assay kit

Transfected lung cancer cell lines were plated on 96-well plates at a density of 5 × 10³ to 1 × 10⁴ per well. Cellular viability was measured by the MTT proliferation assay kit (ATCC, Manassas, VA, USA) according to the manufacturer’s instructions as described in the previous paper from our group59 (link). Each assay was performed in triplicate, and each experiment was repeated at least three times. The extent of cellular survival was represented as a percentage of the first measurement day.

SKOV3, OVCAR8 EIF5A2 KO or OVCAR3 expressing and corresponding control cells (3000/well) were plated into 96-well plates. Cell proliferation was measured at 24, 48 and 72 h using the MTT proliferation assay kit from ATCC (Manassas, VA) according to the instruction of the manufacturer.

Bladder cancer cell lines J82 and SCaBER were plated on a 96-well plate at a density of 5 × 10³ to 1 × 10⁴ per well and incubated overnight at 37°C. The next day, cells were transfected with VGF expression vector (pCMV6-AC-VGF-GFP), empty vector (pCMV6-AC-GFP) (Origene, Rockville, MD) and FuGENE HD transfection reagent (Promega, Fitchburg, WI). Cellular viability was measured by the 3-(4, 5-dimethyl thiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) proliferation assay kit (ATCC, Manassas, VA) according to the manufacturer's instructions after different time points. At the end of each time point, 10 μl of MTT labeling reagent (5 mg/ml MTT) was added to the culture medium, which was then incubated in the dark for a further 4 h at 37°C. This step was followed by cell lysis with the addition of 100 μl of a SDS-based detergent reagent. The plates were incubated for 2 h at 37°C to dissolve formazan crystals. Spectrophotometric readings (570 nm-650 nm) were obtained on a Spectra Max 250 96-well plate reader (Molecular devices, Sunnyvale, CA). Each assay was performed in triplicate, and each experiment was repeated at least three times. Data are represented as the extent of cellular survival expressed as a percentage of control (empty vector).