Three-dimensionally (3D) printed Ti–6Al–4V (Ti64) samples via an electron beam melting (EBM) process were developed to investigate their microstructure and mechanical and tribological properties in comparison with those of commercial Ti64 samples. The 3D-printed Ti64 samples had a heavily twinned and acicular martensitic structure that was responsible for their higher surface hardness than that of the commercial Ti64 samples. The 3D-printed Ti64 samples tested against a 100Cr6 steel counter ball without and with Hank's solution had a higher wear resistance associated with their higher surface hardness than the commercial Ti64 samples. The use of Hank's solution during sliding reduced the wear of the both Ti64 samples as a result of the lubricating effect of the solution. It could be concluded that the 3D-printed Ti64 samples in this study had comparable mechanical and tribological properties to those of the commercial Ti64 samples.