Previous research has demonstrated the effectiveness of applying a simple ageing treatment over the ASTM standard passivation treatment to reduce metal ion release from Ti-6Al-4V alloy into bovine serum solution. The present study aims to further understand the mechanisms behind the improved dissolution resistance exhibited by the aged Ti-6Al-4V alloy using a number of sophisticated surface analysis techniques, X-ray photoelectron spectroscopy and atomic force microscopy. Furthermore, osteoblastic cell culture and functional genomic studies have been employed to fully characterise the biological performance of the treated implants. This study has demonstrated that the passivation treatment results in the elimination of vanadium from the oxide layers. The O/Ti ratio of the passivated sample significantly changes with immersion time in serum, indicating that further oxidation and dissolution processes were occurring. In contrast, the aged oxide surface is more stable and possesses hydroxylated groups, which are thought to play an important role in the reaction with serum proteins. In addition, this research has observed that the side effects of Al₂O₃ grit-blasting and heat treatment associated with the hydroxyapatite plasma-spraying coating cause the change in the Al% surface composition and the loss of chemisorbed H₂O groups; this in turn, affects the metal ion dissolution behaviour of the treated implant. Furthermore, a potentiodynamic test was used to assess the nature of the passive film; the aged surface showed better dissolution resistance and was less affected by bovine serum solution than other treatments. Based on the findings of this study the aged surface has demonstrated much improved dissolution resistance, which is reflected in its biological performance. Moreover, the genocompatibility test is introduced in this study to complement existing biocompatability tests and to provide further detailed information on cell-implant interface reactions.