We have used temperature-sensitive COS cells to design delayed competition experiments in which competition for simian virus 40 (SV40) enhancer factors occurs after enhancer-dependent transcription has been established. The results demonstrate that competition for SV40 enhancer-binding factors has no effect on enhancer-dependent transcription after transcription has been established at the SV40 early promoter. These data show that the enhancer and factors that bind to it are involved in the establishment of stable transcription complexes, although they do not show whether enhancer factors are an integral part of the transcription complexes. Furthermore, the results of delayed competition experiments with a replicating test plasmid are consistent with the possibility that enhancer-dependent stable transcription complexes could be maintained after DNA replication.