Various mammalian cells propagated in serum-free and chemically defined media yielded high titres of Venezuelan equine encephalomyelitis virus. Some difference in maximum titres was noted, depending upon the medium employed. Of the two serum-free media tested, lactalbumin hydrolysate medium was more effective than the chemically defined medium in supporting virus growth. The addition of serum to serum-free cultures at the time of virus inoculation had a pronounced effect characterized by a delay followed by a burst of virus replication to very high titres. Thus, the degree of replication of Venezuelan equine encephalomyelitis virus appeared to be influenced by a variety of unknown nutritional factors.