Effects of growth hormone, insulin and a serum fraction with sulfation factor activity on in vitro incorporation of (3H) leucine into TCA-insoluble protein by diaphragm and costal cartilage of hypophysectomized rats were compared. Tissues were incubated in a medium containing 14 amino acids and glucose. Cartilage was very sensitive to growth hormone in vivo but relatively insensitive in vitro. This tissue was also relatively insensitive to insulin in vitro but very sensitive to sulfation factor. Muscle was less sensitive than cartilage to growth hormone in vivo or sulfation factor in vitro. In all muscle studies combined, the maximum effect of growth hormone in vivo and the maximum effect of either sulfation factor or insulin in vitro each exceeded the maximum effect of growth hormone in high concentration in vitro. Sulfation factor in serum of normal rats was sufficient for stimulation of both muscle and cartilage from hypophysectomized rats in vitro, and the effects were not augmented by a high concentr...