2.6. Assessment of Cell Viability by MTT Assay

HEK293 cells were seeded in a 96-well plate at a density of 30% six hours prior to transfection. Polyethylenimine (PEI) (0.1 mg/mL) was used to co-transfect 15 ng of pCH-9/3091, 15 ng of pCI-neo eGFP, and 85 ng of the left and right TALEN monomer-expressing plasmids. Cells transfected with 170 ng of pUC118 served as the mock transfection control. Cells treated with 50% dimethyl sulfoxide (DMSO) were used as a positive control, and untreated cells served as the negative control. Cell viability was assessed 48 h after transfection. Twenty microliters of 5 mg/mL MTT, made up in PBS, was added to each well and incubated at 37 °C for 1 h. Culture medium was subsequently removed, 200 µL of DMSO added and the cells incubated for a further 5 min. The metabolism of MTT to form blue formazan was determined by measuring the optical densities at 570 nm and 655 nm using an iMARK™ Microplate reader (Bio-Rad, Hercules, CA, USA).