To investigate the natural history of periurethral collagen injection (PCI) over time using serial three-dimensional ultrasonography (3DUS) of the urethra as an objective measure. Retrospective chart review was performed for all patients who underwent PCI between February 1999 and February 2003. All had been diagnosed with stress urinary incontinence due to intrinsic sphincter deficiency without urethral hypermobility, had follow-up data including two or more 3DUS scans within 1 year or more, and had undergone no additional PCIs (ie, remained clinically improved). The follow-up examinations included serial history, symptom and quality-of-life (QOL) questionnaires (Urogenital Distress Inventory, global QOL visual analog scale [scale of 0 to 10, with 0 = best]), physical examination, and 3DUS scans. The primary subjective and objective outcomes were the Urogenital Distress Inventory and QOL scores and the 3DUS-determined periurethral collagen volume and configuration, respectively. Of the 54 patients undergoing PCI during the accrual period, 20 had follow-up of 1 year or longer (mean 2.0, range 1 to 3.8). The mean 3DUS collagen volumes were not statistically different at baseline at a mean of 4 months after PCI (2.9 ± 1.9 cm3) compared with at the last follow-up visit (2.7 ± 1.9 cm3; P = 0.34). The volume retention rate was 97% ± 33% of the baseline volume. The periurethral configuration was circumferential in 80% and asymmetric in 20%, and was maintained over time. The postinjection Urogenital Distress Inventory question 3 (stress urinary incontinence) and QOL scores were significantly improved compared with the pre-PCI evaluations with a mean of 2.4 ± 0.8 versus 1.4 ± 0.7 (P = 0.013) and 7.3 ± 2.6 versus 3.5 ± 2.5 (P = 0.001), respectively. This is the first study to demonstrate the long-term durability of PCI on serial 3DUS in association with improved continence and QOL using questionnaire analysis. This new knowledge provides a technical and therapeutic endpoint for PCI. © 2005 Elsevier Inc.