(1) Background: Due to its high safety potential, one of the most common ADAS technologies is the lane support system (LSS). The main purpose of LSS is to prevent road accidents caused by road departure or entrance in the lane of other vehicles. Such accidents are especially common on rural roads during nighttime. In order for LSS to function properly, road markings should be properly maintained and have an adequate level of visibility. During nighttime, the visibility of road markings is determined by their retroreflectivity. The aim of this study is to investigate how road markings’ retroreflectivity influences the detection quality and the view range of LSS. (2) Methods: An on-road investigation comprising measurements using Mobileye and a dynamic retroreflectometer was conducted on four rural roads in Croatia. (3) Results: The results show that, with the increase of markings’ retroreflection, the detection quality and the range of view of Mobileye increase. Additionally, it was determined that in “ideal” conditions, the minimal value of retroreflection for a minimum level 2 detection should be above 55 mcd/lx/m2 and 88 mcd/lx/m2 for the best detection quality (level 3). The results of this study are valuable to researchers, road authorities and policymakers.