We examined mouse tissue for the expression of macrophage galactose/N-acetylgalactosamine-specific C-type lectin using a rat monoclonal antibody (mAb) specific for this lectin (mAb LOM-14). The binding of mAb LOM-14 was detected in detergent extracts from tissue by means of immunoblotting analysis. It was shown that this mAb did not cross-react with mouse hepatic lectins, a structural homologue. The macrophage lectin was widely distributed among various mouse tissues as judged by the affinity isolation followed by the immunochemical detection. The exceptions were brain, liver, kidney, small intestine, and peripheral blood. Extracts from these organs exhibited, at best, very weak signals upon mAb LOM-14 binding, despite the presence of cells expressing macrophage markers. The most intense signal was observed in the extract from skin, suggesting that cells expressing this lectin are abundant in skin. The tissues shown to contain this lectin were further investigated by immunohistochemical staining of the sections. Cells were distributed in the connective tissue and in the interstice, particularly the dermis and subcutaneous layer of skin. Cells localized in the epithelium of skin (epidermis) or other epithelia that we examined were not stained. Perivascular localization of cells stained with mAb LOM-14 was also demonstrated in cardiac and skeletal muscle tissues. Immunoelectron microscopy revealed the presence of this lectin along the rough endoplasmic reticulum. In conclusion, the distribution of C-type lectin specific for galactose/N-acetylgalactosamine in mice was unique. The connective tissue-specific distribution should provide important information on the biological role of this lectin.