Non-invasive genetic manipulation in primary neurons is important in many areas of neuroscience research. Although highly efficient transfections can be performed using viral methods those procedures come with many drawbacks concerning safety issues. Compared to viral methods, non-viral transfection methods have significantly lower transfection rate which limited its use in neuroscience research. This paper describes a novel microfluidic device that was used to investigate the effect of shear stress on transfection efficiency of lipoplex (DNA entangled with liposome) to primary neurons. The device can be used to simultaneously generate regions with multiple shear stress levels using a single device. This device is compatible with cells growing on a monolayer on a conventional tissue culture Petri dish. When exposed to shear stress, post-mitotic primary rat cortical neurons' transfection rate increased by up to 3-fold when compared to static conventional method. Similar effect was observed with mitotic neuronal cell line NIE-115 where upto 45% transfection efficiency was achieved with the aid of shear stress. Through this research, we demonstrated the efficiency of the reversibly binding microfluidic device in executing transfection experiments and corroborated the fact that shear stress is a new parameter to improve non-viral transfection to cells.