Background: The INTERVAL trial showed that, over a two-year period, inter-donation intervals for whole blood donation can be safely reduced to meet blood shortages. We extended the INTERVAL trial for a further two years to evaluate the longer-term risks and benefits of varying inter-donation intervals, as well as to compare routine versus more intensive reminders to help donors keep appointments. Methods: The INTERVAL trial was a parallel group, pragmatic, randomised trial that recruited participants from 25 centres across England, UK. Here we report on the pre-specified analyses after up to 4 years of follow-up. Participants aged 20-78 years who agreed to continue on their originally allocated inter-donation intervals (men: 12, 10 and 8 weeks; women: 16, 14 and 12 weeks) were block-randomised (1:1) to routine versus more intensive reminders using computer-generated random sequences in blocks of sizes 6 or 8 within the main trial arms. Participants were masked to their reminder allocation. The pre-specified primary outcome was units of blood collected per year (full donation unit=470mL) analysed by intention-to-treat. The trial is registered with ISRCTN, number ISRCTN24760606, and has completed. Findings: Between October 19, 2014, and May 3, 2016, 20,757 blood donors (10,843 men, 9914 women) participated in the extension study (median follow-up 1.1 [IQR 0.7-1.3] years). Compared to routine reminders, more intensive reminders increased blood collection by an average of 0.11 (95% CI: 0.04-0.17) units/year in men and 0.06 (0.01-0.11) units/year in women. During the extension study, each week shorter inter-donation interval increased blood collection by 0.23 (0.21-0.25) units/year in men and 0.14 (0.12-0.15) units/year in women (both P<0.0001). More frequent donation resulted in more deferrals for low haemoglobin, lower mean haemoglobin and lower ferritin concentrations (all P<0.0001). No statistically significant differences were observed in quality of life, serious adverse events, or self-reported symptoms. Compared with the main trial, blood donation rates and mean haemoglobin concentration decreased while symptoms increased during the extension study. Interpretation: During a period of up to four years, shorter inter-donation intervals and more intensive reminders resulted in more blood being collected, but in lower haemoglobin concentrations decreased and in more self-reported symptoms compared to the initial two years. / NHS Blood and Transplant, National Institute for Health Research, UK Medical Research Council, and British Heart Foundation.