We sought to determine the efficacy of remaking initially unsatisfactory cervicovaginal ThinPrep (Cytyc, Boxborough, MA) specimens with and without the addition of glacial acetic acid (GAA) and the effect on human papilloma virus (HPV) Hybrid Capture II (HC2; Digene, Gaithersburg, MD) testing. A total of 583 initially unsatisfactory ThinPrep slide preparations were identified, and remakes were made with the residual in the PreservCyt (Cytyc) vials with (n = 455) or without (n = 128) GAA. Clinical follow-up information was obtained. The addition of GAA resulted in a 56.5% reduction in unsatisfactory cases, compared with a 26.6% reduction without GAA. Neoplasia and atypia were detected in the reprocessed specimens. The addition of GAA resulted in false-positive HC2 test results in 10 of 10 cases. Neutralization of the specimen may reverse this effect. Reprocessing unsatisfactory ThinPrep specimens with GAA can substantially reduce the overall unsatisfactory rate and result in the detection of significant lesions. However, the addition of GAA can result in false-positive results on HC2 HPV tests.