Primary effusion lymphoma (PEL) presents as a serous lymphomatous effusion without tumor masses exclusively in body cavities and mainly occurs in human immunodeficiency virus-1 (HIV-1)-infected patients. We established a new PEL cell line, designated GTO, from the pericardial effusion of a 39-year-old Japanese patient with acquired immunodeficiency syndrome-related PEL. This cell line was infected with human herpesvirus-8, but not with Epstein-Barr virus. Southern blot hybridization demonstrated that GTO cells display monoclonal rearrangement of the IgH gene, suggesting clonal B cell proliferation. GTO cells weakly express or lack T cell-associated markers (CD3, CD5, CD8), the majority of B cell-associated markers (CD19, CD20, CD21, CD79a), the alpha chains of beta 2 integrins (CD11a, CD11b, CD11c), HLA-DR, CD30, and surface immunoglobulin (sIgM, sIgG sIg kappa, sIg lambda), TCR (alpha/beta, gamma delta), but express CD45, and post-germinal center B cell/plasma cell-associated antigens (CD38, CD138). They also express a high level of cell-surface CD4 and can be infected by HIV-1. Immunodeficient mice intraperitoneally xenografted with GTO cells developed ascites containing lymphoma cells. The establishment of GTO and a GTO xenograft mouse model may help to provide insights toward a better understanding of the pathogenesis of PEL and the relationship between HIV-1 and HHV-8.