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Polymerase chain reaction (PCR) and a biotin-labeled DNA probe were used to amplify and detect the genome of infectious bronchitis virus (IBV) from tracheal swabs taken from chickens that were experimentally inoculated with the IBV Beaudette, Arkansas, and Gray strains. The viral genome was successfully detected by PCR and confirmed by dot-hybridization assay using a biotin-labeled DNA probe on days 1, 3, 9, and 14 after exposure. Direct electron microscopy (EM) analysis was used to compare the ability of the two tests to detect IBV from the same tracheal swab samples. The EM analysis did not detect IBV in four of eight necropsy groups that were positive using PCR and the biotin-labeled DNA probe. Although histopathological lesions were observed in the tracheas, no clinical signs or specific antibody response were observed in the birds. The virus was also detected in the allantoic fluid of embryonating chicken eggs that had been inoculated with field samples suspected to be IBV. The field samples were passed four to six times in embryonating eggs, and 10 of 17 samples were positive using PCR and the biotin-labeled probe.