A new B-lymphoma cell line (DEAU-cell line) was established from a diffuse large-cell lymphoma (centroblastic type) and was successfully grafted in athymic nude mice. Monoclonal antibodies (MoAbs) were generated using splenocytes of DEAU-tumor bearing mice. Before the fusion experiments, cellular immunity of the mice bearing growing DEAU tumors was restored by injection of spleen cells from conventional Balb/C mice. Spleen cells from conventional Balb/C mice immunized with DEAU-cell line were also used for the generation of MoAbs. Four MoAbs (DBB.42 and DBA.44 from normal Balb/C mice, and DNA.7 and DND.53 from athymic nude mice) were investigated because they identified B-cell- associated antigens not destroyed by fixatives. DBB.42 recognized a pan- B cell-associated antigen (molecular weight (mol wt) = 45 Kd). DBA.44 detected a B-cell antigen (mol wt not determined) expressed on a subpopulation of B lymphocytes in the mantle zone of lymphoid follicles. DNA.7 also defined a B-cell antigen (43 Kd) mainly expressed on germinal center cells. Similarly, DND.53 recognized a B-cell antigen (two bands of mol wt 20 Kd and 35 Kd, respectively) mainly expressed on germinal center cells and mantle zone lymphocytes and interdigitating reticulum cells in the paracortical area. Major differences were found in the reactivities of these MoAbs on malignant lymphomas. DBB.42 was positive with almost all B-cell lymphomas and some T-cell lymphomas. Within the group of low-grade B-cell lymphomas, DBA.44 reacted principally with hairy-cell leukemia. DNA.7 reacted mainly with high- grade B-cell lymphomas with a weak positivity in low-grade B-cell lymphomas. DND.53 reacted with all but one B-cell lymphoma, cells of histiocytosis X, and Reed-Sternberg cells. These findings indicate that new MoAbs can be generated by using spleen cells from athymic mice bearing human tumors as well as by new lymphoid cell lines. The MoAbs so generated, as in the present study, are deemed potentially useful for the recognition of B-cell lymphomas in routine diagnostic histopathology. In addition, DND.53 could be of value for the diagnosis of histiocytosis X and the detection of Reed-Sternberg cells in Hodgkin's disease.