Using GFP/RFP tagged constructs, easily compare the efficiencies and time-course of expression with different transfection conditions e.g. cationic lipid reagents, amount of DNA, exposure time and presence of serum
Use either T-flasks, microplates or dishes. Low though to high magnification (4x, 10x, 20x). Image multiple areas of your vessel to highlight spatial variations and optimize cell distribution
Prior to transfection, accurately assess cell confluence to standardize cell transfection technique. Post transfection, monitor the effect of reagents on cell health using label-free phase segmentation
Use histograms to monitor the distribution of GFP/RFP intensity of transfected cells
Upper left: Time-course of nuclear GFP expression in A549 human lung carcinoma cells transduced with IncuCyte® NucLight Green lentivirus. Abscissa: time (h), ordinate: fluorescent object count (per mm2). Upper right: IncuCyte® blended HD-phase contrast and green fluorescence image of A549 cells transduced with NucLight Green lentivirus (48h). Note nuclear restriction and homogenous expression of GFP. Transduction of cells with NucLight Green is titratable (lower left: A549 cells, lower right: HUVECs) and transduction efficiencies can be enhanced with Polybrene (lower left: A549 cells).