MTT cell proliferation assay

Proliferation assays were performed using the Cell Proliferation Kit I (MTT) (R&D systems) according to the manufacturer’s protocol. Briefly, HeLa cells were seeded at 2.5 × 10⁴ cells/well in a 96-well plate and transfected with either WT, 3A, or empty vector EV-OSF. Sixteen hours after transfection, cells were treated with inhibitors for 24 h. Each condition was performed in 6 replicates, and measurements were averaged. A total of 10 μL of the MTT reagent was added to each well for 3 h. Then, 100 μL of the solubilization solution was added overnight before measuring absorbance at 570 nm.