A plaque assay system for the study of Herpes T virus in various host cells was examined. Squirrel monkey kidney cells were found to be the most sensitive for plaque assay. The difficulty of obtaining kidney cultures without indigenous viral agents hampered the utilization of this tissue for routine purposes and favored the use of rabbit kidney cells. The optimum temperature for viral adsorption and plaque development was found to be 37°C. There was total absence of plaque development at 25° C. Addition of agar to the infected cultures without washing the monolayers prevented diffusion of virus through the agar, indicating the need for washing cell cultures when used for viral cloning procedures. Plaque yield was influenced by the time of adsorption. Maximum plaque development resulted when the adsorption time was extended to 3 hours. The influence of the inoculum size on plaque titer was paradoxical: the smaller the inoculum size (0.05 ml) the higher the plaque yield. The plating efficiency of Herpes T virus was observed by a linear relationship between virus concentration and plaque numbers. The reproducibility of replicate plaque assay and the ease with which Herpes T plaques developed in most cell cultures studied made this a reliable technique for quantitation purposes.