The pathogenesis of allergic conjunctivitis is largely conjectural. We investigated the possible involvement of respiratory syncytial virus (RSV), a ubiquitous respiratory pathogen, in the development of allergic conjunctivitis through immune mechanisms. A new technique of brush cytology was used to obtain conjunctival cells from 30 patients with allergic conjunctivitis and 20 control subjects. Samples were assayed for the presence of RSV sequences with the reverse-transcription polymerase chain reaction and the nested polymerase chain reaction. Specific primers and inner primers were synthesized on the basis of RNA sequences previously identified. RSV sequences were detected in 7 of 30 (23%) patient samples and 1 of 20 (5%) control samples. Our results proved that polymerase chain reaction could detect RSV sequences in conjunctival samples. RSV may be a significant pathogenic factor in allergic conjunctivitis.