Introduction: This single-centre, retrospective study aimed to assess the efficacy and safety of flexible ureteroscopy (FURS) combined with holmium laser lithotripsy in treating children with upper urinary tract stones. Methods: From June 2014 to October 2015, a total of 100 children (74 boys and 26 girls) with upper urinary tract stones were treated using FURS. A 4.7 Fr double-J stent was placed two weeks before operation. Patients were considered stone-free when the absence of residual fragments was observed on imaging studies. The preoperative, operative, and postoperative data of the patients were retrospectively analyzed. Results: A total of 100 pediatric patients with a mean age of 3.51 +/- 1.82 years underwent 131 FURS and holmium laser lithotripsy. Mean stone diameter was 1.49 +/- 0.92 cm. Average operation time was 30.8 minutes (range 15-60). The laser power was controlled between 18 and 32 VV, and the energy maintained between 0.6 and 0.8 J at any time; laser frequency was controlled between 30 and 40 Hz. Complications were observed in 69 (69.0 %) patients and classified according to the Clavien system. Postoperative hematuria (Clavien I) occurred in 64 (64.0 %) patients. Postoperative urinary tract infection with fever (Clavien II) was observed in 8/113 (7.1%) patients. No ureteral perforation and mucosa avulsion occurred. The overall stone-free rate of single operation was 89/100 (89%). Stone diameter and staghorn calculi were significantly associated with stone-free rate. Conclusions: FURS and holmium laser lithotripsy is effective and safe in treating children with upper urinary tract stones.