In an initial survey of 16 cases of Hodgkin's disease, tissues from one case of nodular sclerosing Hodgkin's disease, a recurrence with numerous Reed-Sternberg cells, demonstrated faint heavy- and light-chain immunoglobulin gene rearrangements. Analysis of seven additional similar cases with extremely numerous Reed-Sternberg cells revealed that six of these seven cases contained clonally rearranged heavy- or light-chain genes. In addition, the original biopsy specimen from the index case (obtained two years prior to the recurrence) had the same pattern of rearrangements of the immunoglobulin genes. In contrast, a germline configuration was observed for the beta T cell receptor gene in all cases. These cases of Hodgkin's disease were also investigated for the presence of Epstein-Barr viral (EBV) genomes by Southern and slot-blot DNA hybridization analysis. Tissues from four of the 21 case studied showed evidence of EBV DNA sequences. Uninvolved lymphoid tissue from two of the positive cases failed to demonstrate viral DNA. To assess clonality of the cells containing the EBV genomes, the tissues positive for EBV DNA were also hybridized with a restriction fragment probe for the terminal sequences of the EBV genome. By this analysis three of the four cases demonstrated a clonal population of EBV-infected cells.