Using lectin-free IL-2 as the only initial stimulus, bulk cultures and T-cell clones were established from synovial fluid (SFL) and peripheral blood lymphocytes (PBL) of a patient with rheumatoid arthritis (RA). The cloning efficiency of growing bulk cultures was 3%-4% as evaluated by Poisson statistics and was not enhanced by the addition of autologous synovial fluid or serum. The majority of the cloned T cells expressed the OKT8+ phenotype; several clones were OKT4+ and one clone expressed OKT8+ and OKT4+ antigens. None of the cloned T cells exhibited high NK or lectin-dependent cytotoxicity, although bulk cultures had high NK activity. In primed lymphocyte typing responses, bulk cultures and two T-cell clones established from rheumatoid SFL and PBL showed consistent autoreactivity, which we have never before observed with MLC-derived bulk cultures and T cell clones. One of the autoreactive rheumatoid T-cell clones (B25) was found to provide strong helper activity to autologous B cells in the absence of mitogen. Attempts to reveal reactivity of RA-derived T-cell clones to microbial antigens have so far only been successful with Mycoplasma pneumoniae preparations. Careful analysis of this reactivity revealed, however, that Mycoplasma pneumoniae induces a stimulator cell-dependent mitogenic effect rather than an antigen-specific MHC-restricted T-cell proliferation.