Background: Kinesin Family Member 3B (KIF3B) is one of the most ubiquitously expressed KIFs, which is related to lots of physiological responses, and KIF3B has been implicated in carcinogenesis such as hepatocellular carcinoma cell. However, the expression of KIF3B has not been researched in pancreatic cancer as well as the clinical significance. Methods: Immunohistochemical assays were performed to detect the expression levels of KIF3B in the tumor tissues and adjacent non-tumor tissues. Patients were sequentially divided into different expression levels of KIF3B group based on the staining intensity of FKIF3B in tumor tissues. The link between KIF3B expression and clinical characteristics were investigated, and the role of KIF3B on pancreatic cancer cell proliferation was detected by colony formation and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, respectively. And the proliferation related proteins such as Ki67 and proliferating cell nuclear antigen (PCNA) were detected by Western blot. The possible effects of KIF3B on tumor growth were assessed in vivo. Results: KIF3B was high expressed in human pancreatic cancer tissues. We also found KIF3B was significantly associated to the pTNM stage (*p = 0.018), lymph node metastasis (*p = 0.040) and vascular invasion (*p = 0.034). We reported that increased expression of KIF3B was significantly correlated with poor clinical outcome in our clinical cohort of pancreatic cancer. Furthermore, functional assays revealed that knockdown KIF3B in vitro and in vivo might inhibit cancer cells proliferation by affecting Ki67 and PCNA. Conclusions: Our data suggested that KIF3B was associated with pancreatic cancer malignant progression especially proliferation. Hence, KIF3B might be served as a potential therapy target of pancreatic cancer in clinical treatment.