Highly ordered TiO2 nanotubes were obtained by anodization on Ti6Al4V substrates manufactured by electron beam powder bed fusion (E-PBF). Effects of anodization parameters such as anodizing time, stirring, fluoride concentration, and water content were analyzed in an organic electrolyte (ethylene glycol) that contains ammonium fluoride. The ordering of the nanotubes was measured by regularity ratio calculations based on fast Fourier transform (FFT) from SEM images. It was found that for the processed specimens, the highest ordering of the TiO2 nanotubes was reached at 30 V for 5000 s with a concentration of 9 vol% H2O and 0.4 wt.% NH4F, exhibiting nanotubes free of delamination, cracks, and coral-like structures with a regularity ratio (RR) of 1.91. This work offers a simple method for creating homogeneous and organized TiO2 nanotubes on Ti6Al4V substrates manufactured by E-PBF which potentially improves its functionality in diverse industrial applications such as nanosensors, controlled-release substances, solar cells, water splitting, electrochromic devices, and Li-ion battery anodes. Graphical Abstract: [Figure not available: see fulltext.].