Purification of the antigenic components of pigeon dropping extract, the responsible agent for cellular immunity in pigeon breeder's disease.

Abstract

Pigeon breeder's disease (PBD) is a lung disease caused by inhalation of antigens derived from pigeons. This study was undertaken to characterize the responsible component of pigeon dropping extract (PDE) for PBD. First, crude PDE was applied to SDS-PAGE followed by immunoblotting by using antibodies in bronchoalveolar lavage (BAL) fluid. Second, 9 bands of PDE were separated by SDS-PAGE and used for antigen-induced PBMCs. Finally, amino-terminal sequencing was conducted on an isolated 21-kd protein by 2-dimensional electrophoresis. Immunoblots with BAL fluid from patients with PBD identified 9 bands. Similar patterns were observed by using BAL fluid from 10 control patients (9 with summer-type hypersensitivity pneumonitis or idiopathic pulmonary fibrosis and 1 asymptomatic breeder), except for the 21-kd protein, which was detected in 10 patients with PBD and 1 asymptomatic breeder. The stimulation indices of PBMCs determined by using proteins electroeluted from the 9 bands were higher in patients with PBD than in the 10 control patients. The 21-kd protein was separated into 5 spots by 2-dimensional electrophoresis; these spots were all reactive with BAL fluid from patients with PBD as determined by immunoblotting. The sequence of the 21-kd protein had 57% identity to a Saccharomyces cerevisiae chromosome X reading frame. A synthetic peptide, derived from the amino acid sequence of the N-terminal of the native protein, induced significant proliferation of PBMCs obtained from 5 patients with PBD, but not with PBMCs obtained from control patients. The 21-kd protein is the only protein that identified individuals exposed to pigeons by immunoblotting. Only PBMCs from patients with PBD showed significant proliferation to the 21-kd protein and to the synthetic peptide on the basis of the N-terminal sequence of the native peptide. The 21-kd protein will be an important antigen for studies on the epidemiology, diagnosis, and pathogenesis of PBD.