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Background It is a challenge for the surgeons to accurately diagnose the pancreatic masses preoperatively,which decides the choice of surgical managements and subsequently results in different survivor outcomes,operative complications,and mortality rates.The purposes of this study were to evaluate the diagnostic role that intra-operative puncture biopsy may play in pancreatic masses and to explore the relevant factors influencing the diagnosis.Methods A retrospective study was performed on 94 in-patients admitted to Peking University First Hospital for pancreatic masses during the period from June 1994 to December 2007.They all underwent intra-operative puncture biopsy during exploratory laparotomy.The sensitivity and specificity of intra-operative puncture biopsy were calculated and the relevant factors to the diagnosis of biopsy were selected for the statistical analysis.Results The overall sensitivity,specificity,positive predictive value,and negative predictive value of intra-operative puncture biopsy were 76.0％,94.7％,98.3％ and 50.0％,respectively.The analysis of bivariate correlations showed that the size of the pancreatic masses (P=-0.000),the number of puncture biopsies (P=0.000),and the presence of pancreatic fibrosis (P=-0.012) had statistic significance for the diagnosis.But the multivariate analysis identified the size of the pancreatic masses (P=0.004) and the number of puncture biopsies (P=0.000) as independent predictive factors for intra-operative puncture biopsy.In addition,as the number of puncture biopsies increased,the sensitivity and specificity of diagnosis was improved (P=0.000).The sensitivity and specificity of intra-operative puncture biopsy were found to be lower for the pancreatic masses less than 25 mm compared with the masses larger than 25 mm (P=0.000).It was noted,however,that even if the masses were less than 25 mm,the sensitivity and specificity could be improved significantly as the number of puncture biopsies reached 3 to 6 (P=0.007).Conclusions Intra-operative puncture biopsy is simple and accurate for qualitatively differentiating various types of pancreatic masses.Three to 4 biopsies could significantly improve the diagnostic effect for pancreatic masses,even if the masses are less than 25 mm in size.