At least three laboratories have reported that the P3HR-1 line, which had originally produced transforming Epstein-Barr virus (EBV), now produces only the nontransforming variant. Studies to determine whether these findings were universal or a consequence of specific cell lines or culture conditions were undertaken in P3HR-1 cultures of identical HLA types from five sources. All of the EBV preparations derived from cell lines cultured at 32, 34, and 35 degrees C transformed cord blood lymphocytes, whereas virus propagated at 37 degrees C did not usually transform. Furthermore, indirect immunofluorescence revealed that a monoclonal antibody directed against transforming EBV membrane glycoprotein bound to 10-12% of the P3HR-1 cells that had been continuously propagated at 34 degrees C, but the antibody did not bind to the same cells cultured at 37 degrees C. Although virus expression was completely repressed in transformed cord blood cells, transforming virus could be rescued by superinfection with nontransforming P3HR-1 EBV. Cells transformed with P3HR-1 virus induced poorly differentiated lymphomas in athymic nude mice after seven or eight passages. Whether all P3HR-1 cells have the potential to produce detectable quantities of transforming virus remains to be determined.