Electronic toll collection (ETC) plays, as part of transport demand management (TDM) measures, an important role in preventing traffic congestion and improving the environmental conditions in urban and rural areas. An attempt is made in the framework of this paper to evaluate the overall performance of a toll station when a lane is dedicated to ETC. The case study refers to a toll station in the Thessaloniki Metropolitan Area, Greece. Scenarios considered specific traffic characteristics, variable toll booth setups, and different penetration rates of the ETC tag users for car and heavy vehicles. The tool used in the evaluation process was the PTV Vissim traffic simulation software. The operation of the toll station during a specific peak-hour period was simulated with the aid of the specific software. In total, 39 alternative scenarios were developed and compared to determine the level of penetration rate for which the ETC lane would be effective for different toll booth setups. Results showed that when the right lane of the toll station is converted to ETC lane, the penetration rate of this lane must be greater the 15% for the private vehicles and 20% for the heavy goods vehicles (HGV) to reduce traffic congestion and to improve environmental conditions. It was also found that when an additional ETC lane was introduced to the existing toll station set up, traffic congestion and the associated environmental conditions were much improved even for low penetration rates. It must be noticed that the results from the use of discounted cash flow methods like internal rate of return (IRR), net present value (NPV) and benefit–cost ratio (BCR) showed that all economic indicators converge as penetration rate increases in all toll booth setups. Therefore, there is a specific penetration rate threshold above which the economic viability of the investment is secured. These findings can assist the design of an effective policy in terms of the optimized operation of a toll station and sustainable mobility planning.