Transfection assays were performed in triplicate in the following manner. HeLa cells were cultured in DMEM supplemented with 10% FBS according to ATCC recommendations. Cells were seeded in 24-well plates at 7.5 x 10 4 cells/well and incubated for 24 h before transfection. Polyplexes were formed by diluting 20 μL of 0.1 μg/μL pGL3 plasmid DNA solution with 80 μL of phosphate-buffered saline (PBS) in a 1.5 mL microcentrifuge tube. Polymer solution (100 μL) at various concentrations was added dropwise to the DNA solution under constant agitation to achieve the desired polymer/DNA weight ratio. Polyplexes were incubated at room temperature for 30 min, after which 200 μL polyplex suspension was deposited into 2.8 mL serum-containing medium. The growth medium was aspirated from cells and replaced with 750 μL of polyplex/growth medium suspension (0.5 μg DNA/well). After 4 h, the transfection medium was removed and replaced with a standard growth medium. Transfection efficiency was quantified via luciferase expression determined at 24 h post initial transfection using a Promega luciferase assay kit. Luminescence was quantified using a Synergy 2 plate-reader and expressed in relative light units (RLU). Results were normalized to total cell protein using a bicinchoninic acid (BCA) assay from G-Biosciences (St. Louis, MO).