Road Traffic Crashes (RTCs) are one of the leading causes of premature deaths in Ghana. Recently, after the rehabilitation of a very critical link on the N1-Highway in Accra, RTCs have soared. The prevalence of traffic injuries and fatalities has attracted the attention of both, road safety professionals and policy makers. Road user behavior has been established to contribute significantly to RTCs. The object of this study was therefore to assess the attitude of both pedestrians and motorists in relation to compliance with traffic safety regulation on the N1-Highway. Covert but unobstructed spot speeds studies were undertaken at accident-prone location on the highway with posted speed limit of 70 km/h. Besides, driver compliance with traffic signal regulations and pedestrian road crossing behavior were also assessed. The 85th percentile speeds realized was 78.2 km/h, which was in excess of the posted speed limit. In all, 42,298 motorists were registered, of which 0.3% was observed run the red traffic lights during the day on the highway. More than 1 in 5 of the 105,151 pedestrian observed jaywalked. Road user behavior undoubtedly, presents a significant road safety challenge on the N1-Highway. Road user education and training must be pursued and sustained, alongside strict enforcement of traffic safety regulations to modify road user, while considering legislation to regulate pedestrian behavior in traffic behavior for safer travels.