Objective To investigate the method and value of tru-cut biopsy（TCB） combined with fine needle aspiration biopsy （FNAB） in the pathological diagnosis of pancreatic carcinoma during operation. Methods From April 2007 to October 2008, 22 cases who were suspected to suffer from pancreatic carcinoma were enrolled into this prospective study. All of them underwent a tru-cut biopsy combined with fine needle aspiration biopsy for the pathological diagnosis during operation. Results Of the 22 patients, 20 were finally diagnosed as having pancreatic carcinoma, while 2 having pancreatitis. The diagnosis of pancreatic carcinoma was confirmed in 19 by tru-cut biopsy combined with fine needle aspiration biopsy, while other 3 cases were not confirmed as pancreatic carcinoma. Among those 3 cases, one was diagnosed as having pancreatic carcinoma with hepatic metastasis by liver nodular biopsy, one as suffering from autoimmune pancreatitis, and another case as having chronic pancreatitis confirmed by follow-up for 9 months without any changes after the operation. The accuracy of FNA, TCB and FNA combined with TCB in the diagnosis for suspected pancreatic cancer were 86.4%, 90.9% , and 95.5% , respectively. No pancreatic fistula and bleeding developed after operation. Conclusion Tru-cut biopsy is more accurate in diagnosis for the suspected pancreatic cancer than fine needle aspiration biopsy during operation. Tru-cut biopsy combined with fine needle aspiration biopsy can improve the accuracy of diagnosis, and is a safe and effective diagnostic method.