This study was designed to determine whether nano-sized ZnO has the potential to cause acute cutaneous irritation using cultured HaCaT keratinocytes and a human skin equivalent as in vitro models, compared to non-nanomaterials. Commercial nano ZnO with different sizes (50 nm and 100 nm) was characterized by dynamic light scattering (DLS) and microscopy (SEM) in different media. Nano ZnO reduced the cell viability of HaCaT in a dose-dependent and time-dependent manner, in a similar way to macro ZnO. However, the 3D-epidermis model revealed no irritation at 1 mg/mL after 24 h of exposure. In conclusion, nano-sized ZnO does not irritate skin, in a similar manner to non-nano ZnO.