Hairy cell leukemia cell lines were established from eight untreated patients using purified B cell growth factor (BCGF) in vitro. These cell lines maintained their original cell surface immunophenotype for about 1 month, after which they began to lose one or more of their characteristic surface antigens. The cell lines also maintained typical hairy cell leukemia morphology for 2-3 months in vitro but later showed an increasing number of multinucleate giant cells that maintained a B cell surface phenotype. The cell lines became independent of exogenously provided BCGF after at least 1 month in vitro and secreted BCGF activity into culture supernatants in most cases. Some cell lines also acquired Epstein-Barr virus nuclear antigen positivity after variable period. Two hairy cell leukemia patients also showed hyperactive T cell responses in vitro and exhibited spontaneous T cell proliferation in culture without exogenously supplied interleukin-2. These T cell lines had the T helper phenotype and secreted significant amounts of T cell-associated lymphokines with BCGF and interleukin-2 activity into culture supernatants.