In the present study, LNA-probe based real-time PCR was designed for the detection and absolute quantification of infectious bronchitis virus (IBV) from the oviduct of unvaccinated and vaccinated hens after IBV challenge. Using a recombinant plasmid standard, the detection limit of the reaction was found to be 10 copies and independent assay runs showed reproducible Ct values. Amongst the unvaccinated hens, the virus could be detected between 6 and 20 days post-infection (p.i.), with a peak of viral load between 10 and 14 days p.i. The virus was also detectable in the oviduct of vaccinated, challenged hens although the viral load was much lower compared to the viral load in the oviduct of unvaccinated, challenged hens. This indicates that rearing phase vaccination can offer significant protection of the fully functional oviduct against a pathogenic strain of IBV. The present test will be useful for the rapid identification of IBV directly from clinical samples. Most vaccination trials investigating the efficacy of vaccines for layer and breeder hens have been conducted based on the respiratory tract response. Evaluation of viral load from the oviduct of vaccinated and unvaccinated hens is an efficient method for assessing oviduct protection in commercial laying hens.