Although the value of cervical cancer screening is widely acknowledged, the effectiveness of an organized cervical cancer screening program in Korea has never been evaluated. We investigated the associations of the frequency of cervical cancer screening with cervical cancer incidence using data from a large prospective cohort study. In this analysis, 253,472 women without a hysterectomy or previous cancer diagnosis were included. Follow-up was between 1995 and 2002. Frequency of Pap smear testing was determined by searching the National Health Examination Database. Using the Korean Central Cancer Registry, 248 cases of invasive cervical cancer and 346 cases of carcinoma in situ (CIS) of the cervix were identified. Subjects screened 2 or more times showed a 71% (corrected reduction 60%) and a 66% (corrected reduction 53%) reduced risk of invasive cervical cancer and CIS of the cervix, respectively, as compared with unscreened subjects [relative risk (RR) = 0.29; 95% confidence interval (CI) = 0.20-0.45; RR = 0.34; 95% CI = 0.25-0.46, respectively]. Women with a normal or benign pap smear had a statistically significantly lower risk of invasive cervical cancer and CIS of cervix compared with those never screened. In age-stratified analyses, there was a significant reduction in cervical cancer incidence among women aged 30 and over who were screened 2 or more times compared with women never screened. The results of this prospective cohort study show that regular screening of cervical cancer reduces invasive cervical cancer incidence and CIS of the cervix among Korean women.