Osteopontin (OPN), a secreted adhesive glycoprotein, is significantly overexpressed in a variety of experimental models of malignancy. Moreover, increased levels of OPN have been detected in the blood of patients with metastatic carcinoma. To investigate OPN expression and distribution in human carcinomas directly, we studied a wide variety of common tumors by Northern analysis, in situ hybridization, and immunohistochemistry. All 14 tumors studied by Northern analysis showed very substantial increases in OPN messenger (m)RNA when compared to corresponding normal tissues. Moreover, intense labeling for OPN mRNA was detected in 71 of 76 carcinomas studied by in situ hybridization. In most of the carcinomas studied (colon, stomach, duodenum, pancreas, breast, lung, bladder, prostate, ovary, thyroid, and melanoma), tumor cells did not label detectably for OPN mRNA; however, macrophages intimately associated with tumor cells labeled strongly for the OPN transcript. In carcinomas of the kidney and endometrium, both tumor cells and host macrophages labeled strongly for OPN mRNA. The presence of OPN mRNA in macrophages was particularly pronounced at the edge of tumors (ie, the tumor/stroma interface) and in areas of tumor necrosis. Although in most cases tumor cells did not label detectably for OPN mRNA, both tumor cells and macrophages stained for OPN protein, suggesting that OPN secreted by macrophages may bind to tumor cells, possibly through the glycine-arginine-glycine-aspartate-serine cell binding domain in OPN. Collectively, these data suggest that OPN functions in adhesive interactions at the tumor/host interface and thereby may influence processes such as invasion and metastasis.