Nih 3 t3 cells

CLL cells (American Type Culture Collection; ATCC; Manassas, VA, USA) were maintained in RPMI medium supplemented with 10% FBS (Lonza Group, Ltd., Basel, Switzerland) and 1% penicillin-streptomycin antibiotic mixture in an atmosphere of 65% humidity with 5% CO₂ and 37°C. The cells were seeded in 96-well plates at a density of 1.2×10⁴ cells per well and incubated for 24 h. The cells were then treated with miRNA-loaded liposomes or blank liposomes (0.1, 1, 10, 50 and 100 µM) and incubated for a further 24 h. The cells were treated for 24 h with increasing concentrations of miRNA-26a (25, 50 and 100 µM) to study concentration-dependent cytotoxicity against leukemia cells. Untreated cells were maintained throughout the study period. The cells were treated with 100 µl 1.25 mg/ml MTT solution and incubated for 4 h at 37°C, after which the formazan crystals were dissolved in DMSO. The absorbance was determined at 570 nm using a microplate reader (Infinite M200 reader; Tecan, Männedorf, Switzerland). For comparison, NIH-3T3 cells (ATCC) was purchased and grown in RPMI medium supplemented with 10% FBS (Lonza Group, Ltd.) and 1% penicillin-streptomycin antibiotic mixture in a humidified atmosphere (65%) with 5% CO₂ and 37°C. The same protocol was followed for MTT assay of these cells.