Introduction of exogenous DNA into mammalian cells represents a powerful approach for manipulating signal transduction. The available techniques, however, are limited by low transduction efficiency and low cell viability after transduction. Here we report a highly efficient molecular delivery technique, named nanotube spearing, based on the penetration of nickel-embedded nanotubes into cell membranes by magnetic field driving. DNA plasmids containing the enhanced green fluorescent protein (EGFP) sequence were immobilized onto the nanotubes, and subsequently speared into targeted cells. We have achieved an unprecedented high transduction efficiency in Bal17 B-lymphoma, ex vivo B cells and primary neurons with high viability after transduction. This technique may provide a powerful tool for highly efficient gene transfer into a variety of cells, especially the hard-to-transfect cells.