Background: Residual kidney function (RKF) has consistently been a predictor of greater survival in maintenance hemodialysis (MHD) patients. The relationship between hemodialysis (HD) treatment frequency and RKF preservation has not been well examined. We hypothesized that initial twice-weekly HD helps in maintaining a longer RKF. Methods: In a dialysis center in Shanghai, 168 ESRD patients were screened and finally 85 patients were identified for this main cohort study. We first examined these 85 MHD patients; 30 of them were initiated with twice-weekly HD for 6 months or longer and 55 patients were started and maintained on thrice-weekly HD treatment. Then a subcohort study in 48 incident MHD patients was implemented to assess the independent risk factors responsible for RKF decline during the first year of HD therapy. Multivariate logistic regression analysis was then employed to examine the odds ratio of RKF loss. Results: The main cohort study showed that the clinical outcomes were almost the same between the two groups in 85 patients, but the percent of patients with RKF loss was significantly lower in the twice-weekly group compared with the thrice-weekly group, especially during the first year of HD initiation. In the 48 incident MHD patients, we found no significant differences between the two groups except for variations in the HD frequency, weekly Kt/V. The multivariate analysis showed that factors such as the male gender, HD frequency, URR and intradialytic hypotension episode were associated with RKF loss, and the odds ratio of RKF loss for each additional HD treatment per week was 7.2. Conclusion: Twice-weekly HD during the first year of dialysis therapy appears to be associated with better RKF preservation.