The mechanisms by which triiodothyronine (T3) stimulates growth and maturation of growth-plate cartilage in vitro were studied by incubating embryonic chick pelvic cartilages in serum-free medium in the presence and absence of T3 for 3 days. To determine whether T3 might stimulate production of somatomedins by the cartilage, medium from cartilage incubated with and without T3 was assayed for somatomedin C (Sm-C) by radioimmunoassay. No difference in Sm-C content was found. However, cartilage incubated with T3 and increasing amounts of human Sm-C (0.5-20 ng/ml) weighed more and had greater amounts of glycosaminoglycan than cartilage incubated in the same concentrations of Sm-C without T3, suggesting that T3 enhances the growth effect of somatomedin. We added a monoclonal antibody to Sm-C (anti-Sm-C) to the organ culture to determine whether T3's stimulatory effect on cartilage growth could be blocked. The anti-Sm-C inhibited growth of cartilage incubated in medium alone and blocked the growth response to T3. By using alkaline phosphatase as a biochemical marker to follow maturation, we found that T3 stimulated a 57% increase in alkaline phosphatase activity above cartilage incubated in medium alone and that anti-Sm-C did not inhibit T3's stimulatory effect on alkaline phosphatase activity. We propose two different mechanisms by which T3 affects growth-plate cartilage: T3 promotes cartilage growth primarily through enhancing the effect of somatomedin, and T3 stimulates cartilage maturation possibly by accelerating the normal process of cartilage differentiation from proliferative to hypertrophic chondrocytes.