Introduction The present study aimed to evaluate the effect of a longer interval between the first and second stages of infected total knee arthroplasty (TKA) revision on the clinical and functional outcome. Methods This study included a total of 56 patients who underwent two-stage revision TKA with a dynamic spacer with a minimum of 2 years of follow-up. Patients were categorized into two groups according to time with the spacer: < 3 months (Group 1, 31 patients) or > 3 months (Group 2, 25 patients). Clinical outcome and quality of life were assessed by knee range of motion (ROM), Knee Society Score for Knee (KSS-K), Knee Society Score for Function (KSS-F) and Short Form 36 (SF-36). Results The mean follow-up period was 48 +/- 19.1 months (range, 24-84 months). The KSS-K, KSS-F, and ROM values were significantly higher in Group 1 than in Group 2 (p < 0.05). The SF-36 scores for general health, physical function, and bodily pain were significantly higher in Group 1 (p < 0.05). Re-infection occurred in 10 patients (17.8%). Time with spacer was not associated with re-infection development (Group 1,n = 6, 19% vs. Group 2,n = 4, 16%;p > 0.05). Conclusion Increased duration with a spacer is associated with poorer clinical and functional outcomes as well as higher treatment costs in two-stage revision knee arthroplasty. Surgeons can attempt to reduce the time patients spend in a spacer to obtain better postoperative functional outcomes, as well as a better quality of life.