[Objective] To investigate the accuracy and efficacy of 3D printed patient-specific instrumentation (PSI) assisted high tibial osteotomy(HTO) in varus knee osteoarthritis(KOA), and to compare with traditional osteotomy. [Method] 35 patients with KOA who underwent medial open wedge HTO in the Department of orthopaedics of our hospital from January 2019 to February 2021 were analyzed retrospectively. They were divided into two groups: 3D group (17 cases) underwent 3D printing PSI assisted osteotomy; The traditional group (18 cases) underwent traditional osteotomy. The perioperative conditions, follow-up and imaging data were compared between the two groups.[Results] The total length of surgical incision, operative time, blood loss, fluoroscopy times, ground walking time and postoperative hospital stay in the 3D group were significantly better than the traditional group, with statistically significant differences (P<0.05). The incidence of early complications in the traditional group was higher than the 3D group, but there was no significant difference (P>0.05). The mean follow-up time was (14.34±2.03) months from 12 to 18 months. The 3D group returned to full weight-bearing activity earlier than the traditional group, but the difference was not statistically significant. VAS score, HSS score and knee extension ROM in 2 groups were significantly improved over time (P<0.05). At 6 and 12 months after surgery, WBLR and MPTA in the 3D group were significantly higher than the traditional group (P<0.05), and the K-L grading of knee osteoarthritis in both groups was significantly improved after operation (P<0.05). [Conclusion] Compared with conventional osteotomy, 3D printed PSI assisted OWHTO has fewer intraoperative fluoroscopy, shorter operative time, more accurate osteotomy, and lower limb line correction is more ideal.