At the cellular level, dynamic strain plays a key role in cell stimulation and organization of the extracellular matrix. Although positive effects of physical strain on tendon tissue are well known, little knowledge exists on how mechanical strain affects tendon cells. In this study, human tendon fibroblasts from patellar tendon were cultured on silicone dishes. Subsequently, cyclic biaxial mechanical strain was applied to the dishes for 15, 30, and 60 minutes using a specially developed cell stretching system. After the fibroblasts were strained, cells were tested for proliferation at 6, 12, and 24 hours. As a control, cells were grown on silicone dishes but did not receive any strain. A biphasic response in proliferation was observed for the 15- and 60-minute strain periods: at 6 hours and 24 hours there was more proliferation than at 12 hours. After a strain duration time of 30 minutes, a lower proliferation rate was measured compared with control levels. This study shows that application of mechanical stress to tendon fibroblasts resulted in an alteration of cellular proliferation depending on the stress time. Our results may implicate future modifications in the treatment of ligament and tendon injuries.