( Cells are stressed In g...)Transfection Reagent Selector Kit Handbook

Cells are stressed In general, avoid stressing cells with temperature shifts and long periods without medium during washing steps. We recommend performing transfection experiments in the presence of serum, so that cells are not deprived of necessary growth factors and nutrients. Vector related influences Toxic effects may arise if a plasmid encoding a toxic protein is used, or if too much plasmid with a high expression rate is used. Conversely, if insufficient plasmid with a low expression rate is used, transfection efficiency may be too low. Optimization of plasmid DNA concentration, as described above and in the optimization section, is recommended for every new plasmid and/or new cell line used. Variable transfection efficiencies in replicate experiments Inconsistent cell confluency in replicate experiments Count cells prior to seeding to ensure that the same number of cells are seeded for each experiment. Keep incubation times between seeding and complex addition consistent between experiments. Possible mycoplasma contamination Mycoplasma contamination influences transfection efficiency. Variations in the growth behavior of mycoplasma-infected cells will lead to different transfection efficiencies between replicate experiments. Cells have been passaged too many times Cells that have been passaged for an extended number of times tend to change their growth behavior, morphology, and transfectability. When cells with high passage numbers are used for replicate experiments
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