Naturally derived tendon scaffolds have the potential to improve the treatment of flexor tendon injuries. Seeded and unseeded tendon scaffolds were maintained in the presence or absence of physiologic strain for 7 days. After 7 days, the tensile properties and associated messenger RNA expression were compared. Seeded scaffolds maintained in the absence of strain had significantly lower tensile properties than unseeded tendons and fresh-frozen tendons. The loss of tensile properties was associated with elevated matrix metalloproteinase-2 and collagen III expression. Tensile properties of seeded scaffolds maintained in the presence of strain for 7 days after seeding did not differ from those of fresh-frozen tendons. This study demonstrates that the tensile properties of seeded, naturally derived tendon scaffolds will degrade rapidly in the absence of cyclic strain. Seeded scaffolds used for tendon reconstruction should be maintained under cyclic strain to maintain essential tensile properties.