Guinea pig hindlimbs were unilaterally immobilized at resting length to evaluate histochemical, biochemical, and contractile properties of immobilized muscle. Contralateral limbs remained unrestrained. Four weeks later contractile properties were measured under chloral hydrate anesthesia. Average time-to-peak tension of the immobilized soleus was 30% less, whereas that of the gastrocnemius was not significantly changed relative to contralateral muscles. Immobilized soleus muscles acquired as much as 25% fibers with high alkaline myofibrillar adenosine triphosphatase activity; these fibers do not occur in the normal muscle. Neither the immobilized soleus nor gastrocnemius fatigued more quickly than their contralateral counterparts. In the immobilized gastrocnemius myofibrillar protein (mg/g muscle) decreased to 76% and maximum tetanic tension to 70% of contralateral values. However, tetanic tension per gram wet muscle weight or 100 mg myofibrillar protein was significantly greater in the immobilized gastrocnemius. No specific factor responsible for the increased tetanic tension could be identified.