C1-hemolytic activity was synthesized and secreted by all nine human fibroblast cell lines examined. The magnitude of the hemolytic titer appeared to be partially dependent on the batch of fetal calf serum used to culture the cells. The fibroblast C1 hemolytic activity was destroyed by heating at 56°C and, after activation, by treatment with 2 mM-DFP. Rabbit antihuman C1q inhibited the fibroblast C1-hemolytic activity. Biosynthesis of fibroblast hemolytic activity was inhibited by the presence of cycloheximide and regained on its removal. Fibroblasts were grown in the presence of radioactive amino acids and the culture media examined for the incorporation of radioactivity into C1q, C1r, and C1s as judged by the formation of specific immunoprecipitates. Polyacrylamide gel electrophoresis in sodium dodecyl sulphate of the immunoprecipitates formed using antisera against C1r and C1s provided convincing physiochemical evidence for the de novo biosynthesis of these subcomponents.