Introduction In this trial, we isolated and cultured pancreatic cancer stem cells (CSCs) to produce a vaccine and prospectively evaluated its safety and efficacy in low-,medium-, and high-dose groups.Material and methods Between February and October 2014, we enrolled 90 patients who met the enrollment criteria and assigned them to three groups (n=30).CSC-specific and CSC-non-specific immunity pre-and post-vaccination were compared by Dunnetts multiple comparison test (one-way ANOVA).The data are presented as the mean±standard deviation.Local and systemic adverse events were recorded in the nursing records and compared using the Chi-square test.All statistical analyses were conducted using GraphPad software (GraphPad, San Diego, CA, USA).Results Throughout the trial, an injection site reaction was the most common reaction (54%), and fever was least common (9%).The incidence of these side effects did not vary among the three groups.When the pre-and post-vaccination immunity was compared, we found that both CSC-nonspecific and CSC-specific responses were significantly increased in the high-dose group.Conclusion This study is the first clinical trial of a pancreatic CSC vaccine and preliminarily proves its safety and efficacy.