In the current study we have compared the effects of heat and radiofrequency plasma glow discharge (RFGD) treatment of a Ti6Al4V alloy on the physico chemical properties of the alloys surface oxide Titanium alloy (Ti6Al4V) disks were passivated alone heated to 600 C or RFGD plasma treated in pure oxygen RFGD treatment did not alter the roughness topography elemental composition or thickness of the alloys surface oxide layer In contrast heat treatment altered oxide topography by creating a pattern of oxide elevations approximately 50-100 nm in diameter These nanostructures exhibited a three fold increase in roughness compared to untreated surfaces when RMS roughness was calculated after applying a spatial high-pass filter with a 200 nm cutoff wavelength Heat treatment also produced a surface enrichment in aluminum and vanadium oxides Both RFGD and heat treatment produced similar increases in oxide wettability Atomic force microscopy (AFM) measurements of metal surface oxide net charge signified by a long-range force of attraction to or repulsion from a (negatively charged) silicon nitride AFM probe were also obtained for all three experimental groups Force measurements showed that the RFGD treated Ti6Al4V samples demonstrated a higher net positive surface charge at pH values below 6 and a higher net negative surface charge at physiological pH (pH values between 7 and 8) compared to control and heat treated samples These findings suggest that RFGD treatment of metallic implant materials can be used to study the role of negatively charged surface oxide functional groups in protein bioactivity osteogenic cell behavior and osseointegration independently of oxide topography Published by Elsevier B V