To evaluate the efficiency with which intra-and extracellular magnetite nanoparticles (MNPs) induce cell death under an alternating magnetic field (AMF), we comparatively investigated two systems consisting of MCF-7 human breast cancer cells and MNPs: one is a "simply-added" system, which is essentially a simple mixture of cells and extracellular MNPs, and the other is a "pre-cultivated" system that contains both intra-and extracellular MNPs. We also evaluated the effect of heating by thermostatic water bath (WB) on cell death in both systems. The degree of cell death was greater under heating with MNPs subjected to AMF than in the WB-heating in both systems. At higher doses of MNPs, the degree of cell death was greater in the "pre-cultivated" system than in the "simply-added" system using both heating methods. The cellular uptake of MNPs induced slight cell damage and caused a high degree of cell death at higher temperatures. A significant decrease in cell viability was observed in the presence of internalized MNPs under the AMF and was suggested to be a result of the combined effect of intracellular heating and cellular damage by MNPs.