Prestressed concrete sleepers (or railroad ties) are designed to carry and transfer the wheel loads from the rails to the track foundation. Over a period of time, a railway track could experience various types of static and dynamic loading conditions, which are attributable to commercial train operations. Previous studies have established two main limit states for the design consideration of concrete sleepers: ultimate limit states under extreme impact and fatigue limit states under repeated probabilistic impact loads. Prestressed concrete has played a significant role in maintaining the high endurance of the sleepers under low to moderate repeated impact loads. Despite the most common use of the prestressed concrete sleepers in railway tracks, their remaining lives are not deeply appreciated nor taken into account for track maintenance and renewal. The present experimental investigation was aimed at determining the residual prestressing force of railway concrete sleepers after revenue services using the dynamic relaxation technique. Fifteen sleepers were extracted from a heavy-haul rail network for testing using experimental facilities at the University of Wollongong in Australia. The structural evaluation program included quasi-static bending tests, dynamic impact tests, and tests to establish the current level of prestress in the steel wires using the dynamic relaxation technique. Two of the sleepers were evaluated for the level of prestressing forces in accordance with Australian standards. It was found that the level of prestress determined using the dynamic relaxation technique was significantly lower than that expected from the theoretical analysis of time-dependent prestressing losses for the concrete sleepers.