NO2 and O3 concentrations were monitored in four one-way road tunnels of the newly constructed Attiki Odos in the area of Athens, Greece using Radiello diffusive samplers. Differences among urban, street-level and tunnel concentration for both NO2 and O3 indicated the vital impact of traffic emissions. NO2 levels grew along the tunnel (from 30.2 ± 3.1 μg/m3) resulting in high concentrations near the exit (up to 90 μg/m3), because of the poor ventilation and the piston-effect of traffic movement. Conversely, O3 concentrations dropped from 39.2 ± 1.0 μg/m3 to almost 1.5 μg/m3, demonstrating the substantial effect of elevated traffic emissions on chemistry and oxidative content of tunnel air. Despite different traffic flows and tunnel length, regression analysis showed that NO2 concentrations were proportionally related to the distance from the tunnel entry; whereas, a rather poor relationship was observed for ozone. The results of this study indicated that the length of the tunnel, a tunnel-design parameter, could be used as a surrogate to monitor air quality even in short tunnels. In addition, passive samplers could be employed to measure the concentrations of NO2 and O3, even in extremely polluted environments such as road tunnels.