The objective of this study was to compare the detection of human papillomavirus (HPV) DNA in women who stopped smoking, and those who continued to smoke. Women entering a smoking cessation programme had tampon specimens collected and analyzed for HPV DNA using polymerase chain reaction (PCR) (L1 consensus primers). Women with HPV DNA in their initial specimens had follow-up specimens collected 1 year later. The stored initial specimens were reanalyzed in the same experiment with the follow-up specimen. In addition, Hybrid Capture and quantitative PCR were used to determine if the change in HPV DNA over the year of follow-up was greater in either group. Of 197 women who were recruited into the study, 57 (29%) were positive for HPV DNA. Second specimens were obtained from 49 of these 57 women. Of these 49 women, 13 had stopped smoking and 36 continued to smoke. The detection of HPV DNA in follow-up specimens was not significantly different in the women who stopped smoking (54%) compared to those who continued to smoke (50%, p = 0.81). Furthermore, the change in the amount of HPV DNA during follow-up was not different in women who stopped smoking compared to women who continued to smoke (p > 0.25). This study suggests that smoking cessation does not affect the detection of genital HPV DNA. On the basis of these findings it is likely that smoking does not increase the risk of invasive cervical cancer by prolonging the detection of HPV DNA.