The simple and efficient separation of histidine-tagged (His-tagged) proteins using nickel ferrite (NiFe(2)O(4)) nanoparticle clusters (NPCs) is described in this article. While nanostructured materials containing Ni(2+) ions for efficient separation were generally synthesized via complex synthetic procedures, the NiFe(2)O(4) nanoparticle clusters used in this work were simply synthesized by a one-pot hydrothermal polyol process. Their large surface area (105.0 m(2) g(-1)) and pore volume (0.32 cm(3) g(-1)) provide sufficient sites for the separation of His-tagged proteins. The high magnetic saturation value (41.3 emu g(-1)) and superparamagnetic property of these nanoparticle clusters lead to a more efficient magnetically recyclable separation of His-tagged proteins. We confirmed that the binding capacity and selective separation ability seen in the first separation were strongly maintained for up to 7 cycles.