An accurate assessment of the driving comfort and safety of road vehicles moving on a long-span bridge subjected to vortex-induced vibration (VIV) is essential for bridge administrators to decide whether the bridge should be closed to traffic. However, previous assessments often ignore the aerodynamic interference between the road vehicles and the bridge deck subjected to VIV. In this study, a specific wind tunnel model is developed to explore the aerodynamic interference between road vehicles and twin-box bridge deck during VIV. The vortex-induced force (VIF) and vortex-induced response (VIR) of the twin-box bridge deck and the aerodynamic forces on the vehicle were simultaneously measured. The influence of the vehicles on the VIV of the deck was investigated, and the influence of the deck vibration on the aerodynamic forces of the vehicle was also explored. The results show that the VIR and VIF of the bridge deck were generally reduced, depending on the type, position, and number of vehicles. The aerodynamic forces of vehicles could be amplified due to the deck vibration. These findings supplement the database of vehicle aerodynamic coefficients for assessing the driving comfort and safety of road vehicles moving on a long-span bridge subjected to VIV.