Protocol for STO Cell Transfection by FuGENE HD
STO cells were seeded the day before transfection with the density 15,000 cells per well in 100 μl complete growth medium DMEM+10% Fetal Bovine Serum.
Prepare 0.02μg/μl pCMVβ plasmid DNA solution in OptiMEM®.
Add 6μl of reagent to 100 μl of OptiMEM® /DNA solution.
Mix carefully by pipetting (10-15 times).
Incubate 5 min at room temperature.
Add 5μl complex per well to the cells, and mix thoroughly.
Place the cells into CO2 incubator for 26-28 hours.
Remove the medium from the well and wash the cells once with 100μl per well PBS.
Fix the cells in the well with 50μl solution of 4% formaldehyde in PBS for 5min at room temperature.
Wash each well with 100μl PBS.
(1/2) Add 50μl per well of substrate/stain solution and incubate the plate overnight at 37°C.
Observe the cells under microscope and evaluate the proportion of blue (β-gal-positive) cells.
Wash each well with 100μl PBS.
(2/2)
