Background and study aim: Typically, pancreatic patient-derived tumor xenografts (PDXs) are established by transplanting large tumor biopsies obtained through invasive surgery approaches into immunocompromised mice. We aimed to develop pancreatic PDXs by transplanting tumor tissue acquired by endoscopic ultrasound (EUS)-guided fine needle biopsies (FNB), assess take rates compared to surgery-derived PDXs, and demonstrate the histological and genetic resemblance to the original tumor. Patients and methods: Biopsies of untreated pancreatic carcinoma were collected at surgery and during EUS and processed to generate PDXs. Results: By centrifugation of FNB-derived tissue prior to engraftment, we achieved an engraftment rate of 60 % (6/10). Despite a decrease in stromal tissue, the general morphology of FNB-derived PDXs was conserved as assessed by histopathology. At the genetic level, somatic mutation and copy number profiles were largely similar to the primary tumor. Conclusion: We show that it is technically feasible to establish pancreatic PDXs using a minimally invasive sampling technique, such as EUS-FNB. Although only a limited amount of tumor tissue was acquired, we obtained results similar to those from surgery-derived PDXs.