A stainless steel substrate with highly ordered self-organized nanopores was decorated with gold nano-structures (Au/NPSS) for use in electrochemical biosensors. The nanopores were formed on stainlesssteel surface using anodization process, filled with Cu using pulsed electrodeposition and followed bygalvanic replacement of Cu with gold. The characteristics of the electrode and its potential application forelectrochemical determination of dopamine (DA) were investigated using scanning electron microscopy,energy-dispersive X-ray spectroscopy, cyclic voltammetry and electrochemical impedance spectroscopy.The results indicated that compared to a smooth gold electrode, the nanostructured gold film formed onthe NPSS has a great active surface area and displays a higher peak current for DA oxidation. Cyclicvoltammetric determination of DA exhibited two wide linear ranges of 3.0–8.0 and 8.0–2000 micro-M with adetection limit of 0.07 micro-M (3Sb/m). Moreover, two lines in the range of 1.0–80.0 micro-M of DA with a limitof detection of 0.02 micro-M (3Sb/m) were obtained using differential pulse voltammetry. Au/NPSS exhibitedgood repeatability and reproducibility, long-term stability and acceptable selectivity. This work opens upnew prospects for the fabrication of inexpensive noble metal-based catalysts supported on NPSS, whichhave a potential application in biosensors