Fresh tumor cells from pleural effusion of a patient with Hodgkin's disease were analyzed cytogenetically, immunologically and enzymocytochemically. They were characterized by the presence of alpha-naphthyl butyrate esterase activity, Fc gamma-receptor, HLA-DR antigen and No. 9 antigen which has been shown to be present in Hodgkin's cells and granulocytes, and the absence of definite T-, B- and myeloid cell markers. The karyotype analysis of these tumor cells revealed chromosome instability, but the clonality was confirmed by the many common abnormalities such as -4, -6, -10, -12, -13, -14, +21, del(X) (q22,q26), del(7) (q32q36), and +der(19)t(19;?). In addition, there were more duplicated tetraploid clones than near-diploid clones. The karyotype of the near-diploid clone was interpreted as: 48, X, del(X) (q22q26), -4, -6, -10, -12, -13, -14, +20, +21, +der(4)t(4;?) (p16;?), del(7) (q32q36), +der(19)t(19;?) (p13;?), +mar1, +mar2, +mar3. The karyotype abnormalities characteristic of lymphomas or leukemias were not found. These results indicate that the tumor cells are not of lymphoid or myeloid lineage. Further studies are needed to determine the cellular origin of the tumor cells of Hodgkin's disease.