Electrospray technique has received increasing attentions for intracellular gene delivery as well as production of nanoparticles. In this study, chitosan/pDNA nanoparticles with N/P ratio of 5 were prepared and transferred to HEK293T cells by electrospray technique. Physicochemical characterization of prepared nanoparticles, including size, zeta potential and entrapment efficiency was performed and attachment of pDNA to chitosan was confirmed by gel agarose electrophoresis. Moreover, transfection efficiency was investigated using flow cytometry. MTT assay was performed for cell viability studies. Nanoparticles were prepared at three pDNA concentrations of 10, 55 and 100 μg/ml in fixed N/P ratio. Size of nanoparticles was obtained as 110, 188 and 240 nm, using DLS. SEM showed size of 102.34 ± 10.66 nm for samples having 55 μg/ml pDNA. Zeta potential and entrapment efficiency were +25 mv and 85±4%m respectively. The effect of pDNA concentration, electrospray time and incubation time on transfection efficiency was investigated using Box-Behnken design. Percent of GFP-positive cells was 41.05 ± 3.04% which was taken as an indicator of transfection efficiency. Transfection efficiency of this method was then compared with that of calcium phosphate (31.1 ± 2.4%), showing improved efficiency. Considering the fact that electrospray is an easy, low cost, one-step process which makes low damage to cells and produces monodispersed nanoparticles, the method is introduced as a fascinating approach in gene transfection.
Enhanced gene transfection efficiency by low-dose 25 kDa polyethylenimine by the assistance of 1.8 kDa polyethylenimine
