Obliterative bronchiolitis (OB) occurs in 50% of patients 2 years after lung transplantation. Although a correlation between OB and indirect recognition of donor peptides has been reported, the relative roles of direct vs. indirect recognition have not been investigated. Limiting dilution analysis was used to determine the frequencies of recipient T-helper cells recognizing donor and third-party alloantigens in 19 patients (8 OB positive, 11 OB negative). The assays were designed to distinguish between indirect and direct presentation. In three patients the direct and indirect assay were performed on the same blood sample. Six out of seven patients with OB were hyperresponsive in the indirect pathway to donor antigens compared to third-party, the corresponding figure for OB negative patients being 2/7. In contrast, 5/7 patients were hyporesponsive in the direct pathway; hyporesponsiveness in the direct pathway did not correlate with freedom from OB. The patients in whom the assays were performed from the same blood sample confirmed that donor specific hyperresponsiveness in the indirect route can coexist with donor-specific hyporesponsiveness in the direct route. In conclusion, lung allograft recipients, like recipients of other organ allografts, become hyporesponsive in the direct route but sensitization via the indirect pathway is associated with chronic rejection.