Based on the physical method of electroporation, nucleofection uses a combination of electrical parameters, generated by a device called Nucleofector, with cell-type specific reagents. The substrate is transferred directly into the cell nucleus and the cytoplasm. In contrast, other commonly used non-viral transfection methods rely on cell division for the transfer of DNA into the nucleus. Thus, nucleofection provides the ability to transfect even non-dividing cells, such as neuron and resting blood cells. Before the introduction of the Nucleofector Technology, efficient gene transfer into primary cells had been restricted to the use of viral vectors, which typically involve disadvantages such as safety risks, lack of reliability, and high cost. The non-viral gene transfer methods available were not suitable for the efficient transfection of primary cells. Non-viral delivery methods may require cell division for completion of transfection, since the DNA enters the nucleus during breakdown of the nuclear envelope upon cell division or by a specific localization sequence.
Optimal nucleofection conditions depend upon the individual cell type, not on the substrate being transfected. This means that identical conditions are used for the nucleofection of DNA, RNA, siRNAs, shRNAs, mRNAs and pre-mRNAs, BACs, peptides, morpholinos, PNA, or other biologically active molecules.