Nonviral gene delivery strategies may show reduced immunogenicity and oncogenicity in comparison to viral vectors. However, this has generally been at the cost of endosomal release and transfection efficiency. In this study, we used poly (N,N-dimethylaminoethyl methacrylate (DMAEMA) / 2-hydroxyethyl methacrylate (HEMA)) nanoparticles as gene carriers due to their biocompatibility and ability to swell at low pH. Once internalized via the endocytosis pathway, the nanoparticles undergo osmotic swelling facilitating endosomal escape of DNA to the cytoplasm. Monodispersed nanoparticles were synthesized by O/W emulsion followed by polymerization. Particles having 200 nm average diameters were characterized by transmission electron microscopy, dynamic light scattering, electrophoresis, and cytotoxicity. For intracellular tracking, particles were conjugated with quantum dots and examined by confocal microscopy at different time points. To examine the efficacy of DMAEMA/HEMA nanoparticles in facilitating endosomal escape of pDNA encoded EGFP, HeLa cells were incubated with DNA-loaded nanoparticles for 24 and 48 h. We demonstrate that high swelling ratios are achieved at low pH, low crosslinking density, and high content of DMAEMA. After 48 h incubation, the transfection efficiency obtained by using DNA-loaded DMAEMA/HEMA nanoparticles was 45 ± 4.8%, in comparison to 40 ± 3.1% for PEI/DNA complexes with a reduction in cytotoxicity of 93 ± 6.1%. Our results show that DMAEMA/HEMA nanoparticles lead to high gene expression which is comparable to the efficacy using polyethyleneimine (PEI)/DNA complexes without the drawback of cell death.