To investigate the legibility and visual comfort of LED traffic signs, an ergonomic experiment is performed on four custom-designed LED traffic signs, including three self-luminous ones as LED lightbox, LED backlight and regional LED backlight, and one non-self-luminous sign with external LED lighting. The four signs are hanged side-by-side and evaluated by observers through questionnaires. The signage dimension is one-sixth of the real freeway traffic signs, and the observation distance is 25 m. The luminance of three self-luminous signs is 216 cd/m2. The illuminance of external LED lighting is 400 lux on the traffic sign. The ambient illuminance is 2.8 and 6.0 lux in two rounds. The results show that self-luminous traffic signs provide superior legibility, visual comfort and user preference than the non-self-luminous one. Among the three self-luminous signs, regional LED backlight is most susceptible to the ambient illumination. LED lightbox has significantly better preference score than LED backlight under darker ambient lighting. Only LED lightbox has significantly better visual comfort than external LED lighting in the brighter environment. Based on the four LED traffic signs evaluated in this study, we suggest LED lightbox as the prior choice. Further investigations on the effect of ambient illumination and other designs of self-luminous traffic signs are in progress.