We studied SPan-1 antigen in 64 patients with pancreatic cancer by measuring serum concentrations with RIABEAD and examining the intratumor staining patterns immunohistochemically. Serum concentrations were elevated (greater than 30 U/ml) in 46 patients (72.0%), roughly the same percentage as with elevated serum CA19-9 concentrations. Ten out of 19 patients without CA19-9 antigen expressed SPan-1, and the combined rate of positivity was 86% (55/64). Immunohistochemically, CA19-9 was localized to the apical surface and the supranuclear cytoplasm of normal epithelial cells, but SPan-1 was localized to the supranuclear cytoplasm or throughout the cytoplasm. After malignant degeneration, both antigens were found over the entire plasma membrane, throughout the cytoplasm (loss of polar distribution) and in the stroma surrounding the cells. SPan-1 was detected in 48 out of 54 adeno-carcinomas of the pancreas (89%), and all cases with an elevated serum concentration displayed stromal staining in the cancer tissues. The utility of SPan-1 antigen as a tumor marker for diagnosing pancreatic cancer was found to be equal to that of CA19-9.