Trypsin enhancement of parainfluenza-3 virus infectivity in HEp-2 cells was demonstrated. A substantial enhancement of infectivity was observed when cell cultures were exposed to trypsin during the adsorption period or when subcultured with trypsin late in the eclipse phase. Time-course studies on virus development indicated that treatment of cells with puromycin or cycloheximide for 1 h postinfection markedly inhibited the synthesis of infectious virus. However, this inhibitory effect could be reversed by the removal of drug and subsequent subculture of trypsinized cells.