Objective: To evaluate the clinical value of nanoknife ablation in treatment of locally advanced unresecable pancreatic cancer. Methods: From September 2015 to March 2016, 18 patients with locally advanced unresectable pancreatic carcinoma received nanoknife ablation treatment. The clinical variables of the patients before and after treatment were comparatively analyzed. Results: All patients underwent successful operation, and all vital signs were stable throughout the procedures. The serum amylase concentration of the patients was significantly increased on postoperative day 1 (P<0.05), but then, it gradually decreased to the normal level; the CA19-9 level of the patients presented a slowly decreasing trend after operation, which was significantly lower than that of preoperative value at each studied postoperative time point (all P<0.05); the myocardial enzyme indexes showed no significant difference before and after operation (all P>0.05). Within postoperative 3 months, pancreatitis occurred in 2 cases (11%), and hypoglycemia, hyperglycemia and gastric emptying dysfunction occurred in one case each (5%), which were all quickly relieved after symptomatic treatment, while no treatment-related complications such as pancreatic leakage, hemorrhage, and bile leakage occurred. The scores for quality of life and Karnofsky of the patients were significantly increased after operation compared with preoperative values (all P<0.05). According to the EORTC criteria, complete metabolic remission was achieved in 2 cases (11%), partial metabolic response was obtained in 10 cases (55%), stable metabolic disease was seen in 3 cases (17%), and progressive metabolic disease was found in 3 cases (17%), and the objective response rate was 67% and disease control rate was 83%, respectively. Conclusion: Nanoknife ablation has high safety and demonstrable efficacy in treatment of locally advanced unresecable pancreatic cancer, and it may have a great application potential.