Despite morphological similarities between adenocarcinomas of the small and the large intestine, recent evidence suggests that both tumor types follow different genetic pathways. In particular, inactivation of the APC tumor suppressor gene, a characteristic alteration of colorectal carcinomas, does not seem to play a significant role in sporadic small intestinal tumorigenesis. We could recently show that inactivating mutations of the SMAD4 gene frequently occur in small intestinal adenocarcinomas. To further elucidate the role of SMAD4 dysfunction for tumor development in the small intestine, we immunohistochemically analyzed 20 sporadic, non-ampullary carcinomas for the expression of the SMAD4 protein. We further determined homozygous SMAD4 gene deletions by real time PCR and compared SMAD4 immunohistochemical data with SMAD4 genetic data. Immunohistochemistry was negative for the tumor cells in two (10%) cases and strongly reduced in four (20%). Negative immunohistochemical staining corresponded with homozygous gene deletions. A regular or only slightly reduced staining pattern was noted in 14 carcinomas, including four tumors with previously identified SMAD4 missense and frame shift mutations. In conclusion, our data suggest a significant role of impaired SMAD4 function in the pathogenesis of small intestinal adenocarcinomas. Furthermore, our results show that SMAD4 immunohistochemistry may serve as a surrogate for analysis of homozygous gene deletions. However, the method fails to identify SMAD4 inactivation due to missense mutations.