Abstract This work deals with surface modification of Ti6Al4V alloy. Compared are the morphology, structure, adhesion and bioactivity of Ti6Al4V after chemical treatment in alkali solution, anodic oxidation in mixed solution of NH4F + H3PO4 and thermal treatment. In both cases of chemical and electrochemical treatment an amorphous layer of TiOx is formed. Results showed that the alkali treatment leads to formation of a 1?m thick bioactive layer with net-structure, which offers a good basis for the growth of Ca-P compounds. The in vitro test confirmed a good bioactivity of this layer. The thermal treatment caused dehydration of this layer, what did not affect the good bioactivity as confirmed by the in vitro test. Anodic oxidation forms a nanotubular layer, which is not bioactive, but the thermal treatment activated this layer.