Introduction: Pancreatic ductal adenocarcinoma (PDAC) remains a major cause of cancer-related death. Since significant upregulation of alpha v beta 6 integrin has been reported in PDAC, this integrin is a promising target for PDAC detection. In this study, we aimed to develop a radioiodinated probe for the imaging of alpha v beta 6 integrin-positive PDAC with single-photon emission computed tomography (SPECT). Methods: Four peptide probes were synthesized and screened by competitive and saturation binding assays using 2 PDAC cell lines (AsPC-1, alpha v beta 6 integrin-positive; MIA PaCa-2, alpha v beta 6 integrin-negative). The probe showing the best affinity was used to study the biodistribution assay, an in vivo blocking study, and SPECT imaging using tumor bearing mice. Autoradiography and immunohistochemical analysis were also performed. Results: Among the 4 probes examined in this study, I-125-IFMDV2 showed the highest affinity for alpha v beta 6 integrin expressed in AsPC-1 cells and no affinity for MIA PaCa-2 cells. The accumulation of 125I-IFMDV2 in the AsPC-1 xenograft was 3-5 times greater than that in the MIA PaCa-2 xenograft, consistent with the expression of alpha v beta 6 integrin in each xenograft, and confirmed by immunohistochemistry. Pretreatment with excess amounts of A2OFMDV2 significantly blocked the accumulation of (125)1-IFMDV2 in the AsPC1 xenograft, but not in the MIA PaCa-2 xenograft. Furthermore, I-123-IFMDV2 enabled clear visualization of the AsPC-1 xenograft. Conclusion: 123I-IFMDV2 is a potential SPECT probe for the imaging of alpha v beta 6 integrin in PDAC. (C) 2014 Elsevier Inc. All rights reserved.