Background Venipuncture (phlebotomy) is an obstacle to subject recruitment and ongoing participation in cohort studies investigating human papillomavirus (HPV) infection. Anti-HPV antibodies are not only detected in serum but also in oral fluid. Objectives To evaluate if oral fluid specimens can be used in lieu of blood specimens for determining HPV antibody status. Study design One hundred and seven paired oral fluid and blood specimens from female university students were tested in a HPV16 ELISA and compared to sexual history and serial genital HPV16 DNA status. Results ELISA results were in agreement in 97% (104/107) of paired sera and oral fluid. Of six women with positive anti-HPV16 serum samples, only three had positive oral fluid specimens. However, the specificity of the oral fluid test was 100% compared to the blood test. Conclusions Detection of antibodies in oral fluid correlated with antibodies but was less sensitive than sera. A larger validation study is required to fully characterize the oral fluid assay.