Recently, major achievements in creating decellularized whole tissue scaffolds have drawn considerable attention to decellularization as a promising approach for tissue engineering. Decellularized tissues are expected to have mechanical strength and structure similar to the native tissues from which they are derived. However, numerous studies have shown that mechanical properties change after decellularization. Often, tissue structure is observed by histology and electron microscopy, but the structural alterations that may have occurred are not always evident. Here, a variety of techniques were used to investigate changes in tissue structure and relate them to altered mechanical behavior in decellularized rabbit carotid arteries. Histology and scanning electron microscopy revealed that major extracellular matrix components were preserved and fibers appeared intact, although collagen appeared looser and less crimped after decellularization. Transmission electron microscopy confirmed the presence of proteoglycans (PG), but there was decreased PG density and increased spacing between collagen fibrils. Mechanical testing and opening angle measurements showed that decellularized arteries had significantly increased stiffness, decreased extensibility and decreased residual stress compared with native arteries. Small-angle light scattering revealed that fibers had increased mobility and that structural integrity was compromised in decellularized arteries. Taken together, these studies revealed structural alterations that could be related to changes in mechanical properties. Further studies are warranted to determine the specific effects of different decellularization methods on the structure and performance of decellularized arteries used as vascular grafts.