The virion polypeptide composition of 3 independently isolated tree shrew herpesviruses (THV) was analyzed by sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis (SDS-PAGE) and by a 2-dimensional technique using isoelectric focusing. Two of the virus isolates analyzed were from malignant tumors; the other isolate (THV, strain 1) was from an apparently healthy animal. The polypeptide patterns of the 3 purified Tupaia herpesvirus isolates were remarkably similar, each consisting of at least 35 polypeptides ranging in MW from 12,000 to 230,000. While the majority of analogous polypeptides of the 3 viruses were of indistinguishable electrophoretic mobility, some (polypeptides of 82,000-86,000) showed small differences in apparent MW which were characteristic of the virus strain. Comparative SDS-PAGE made it possible to distinguish the Tupaia herpesvirus isolates from each other. At least 5 glycoproteins were found in purified THV virions. The 2-dimensional electropherograms revealed at least 47 discernible protein spots, some of which were specific for a given THV isolate and which were detectable even in lysates of THV-infected Tupaia embryo fibroblast cells.