Cracking damage influences the stiffness of the girders. Many articles in the literatures have studied the development of stiffness of the scale-down model; however, full-scale model testing cannot be completely replaced by scale-down testing because of material component characteristics and boundary effects. This paper deals with the effects of cracking damage on the structural static and dynamic stiffness based on three prestressed concrete (PC) girders which were removed from an old bridge. First, the equivalent flexural rigidity of cracked prestressed concrete girder was assessed using the measured load-deflection response under cycles of loading and unloading. Then, after unloading, the frequencies were measured on the PC girders supported by the elastomeric bearings. Next, the development of frequency under different damage was studied, and finally, the dynamic stiffness of PC girders with cracks was assessed. The results indicate that the first frequency is more sensitive to the cracking of concrete compared with the second frequency and that the mode shapes are not sensitive to girder damage. The test girders cannot be simplified as an ideal simply supported beam for the purpose of identifying frequencies. In addition, the “final” (the end of the ultimate load case) equivalent flexural rigidity of the girders is 30% of the “initial” (the beginning of the first load case) equivalent flexural rigidity, compared with 50% in the scale-down test; and the final dynamic stiffness is approximately 84% of the initial dynamic stiffness, whereas the scale-down test is 72%.