The cellular immune response to Borrelia burgdorferi was studied in 24 patients with seropositive and seronegative Lyme borreliosis, 30 patients with arthritides of different origin (non-Lyme arthritides), and 20 normal blood donors. By far, the strongest T cell stimulation was induced by incubation with autologous serum; there was a significantly lower response or no response after incubation with allogeneic or heterologous sera. In patients with Lyme borreliosis, including seronegative patients, there was a strikingly elevated proliferation in response to whole B burgdorferi bacteria (mean 64,750 dpm) compared with that of normal donors (mean 19,700 dpm; P less than 0.0001) and especially that of non-Lyme arthritis patients (mean 11,600 dpm; P less than 0.0001). Levels of proliferation declined significantly in patients with Lyme borreliosis after successful antibiotic treatment. Parallel cultures using B burgdorferi and Treponema phagedenis as antigens showed that cells from patients with Lyme borreliosis responded significantly more to B burgdorferi than to T phagedenis, but this did not occur with cells from individuals with non-Lyme arthritides. There was no correlation between disease stages and proliferation values. These data indicate that lymphocyte proliferation assays may provide an important tool for the diagnosis of Lyme borreliosis, most notably in patients with arthritides and in those who are seronegative. Conversely, the lack of reactivity appears to be a strong indicator of the absence of active Lyme disease. It seems to be crucial, however, to use autologous sera in these assays.