PURPOSE: This retrospective cohort study was conducted to estimate the optimal interval for gastric cancer screening in Korean adults with initial negative screening results. MATERIALS AND METHODS: This study consisted of voluntary Korean screenees aged 40 to 69 years who underwent subsequent screening gastroscopies after testing negative in the baseline screening performed between January 2007 and December 2011. A new case was defined as the presence of gastric cancer cells in biopsy specimens obtained upon gastroscopy. The follow-up periods were calculated during the months between the date of baseline screening gastroscopy and positive findings upon subsequent screenings, stratified by sex and age group. The mean sojourn time (MST) for determining the screening interval was estimated using the prevalence/incidence ratio. RESULTS: Of the 293,520 voluntary screenees for the gastric cancer screening program, 91,850 (31.29%) underwent subsequent screening gastroscopies between January 2007 and December 2011. The MSTs in men and women were 21.67 months (95% confidence intervals [CI], 17.64 to 26.88 months) and 15.14 months (95% CI, 9.44 to 25.85 months), respectively. CONCLUSION: These findings suggest that the optimal interval for subsequent gastric screening in both men and women is 24 months, supporting the 2-year interval recommended by the nationwide gastric cancer screening program.