The localization of immunoreactive retinol-binding protein (RBP) in rat liver was studied by immunofluorescence microscopy. The study employed specific antisera to rat RBP prepared in a rabbit and in a sheep. The indirect, two-stage method of localizing tissue antigens was employed, and livers of both normal and vitamin A-deficient rats were examined. Fab' fragments of immunoglobulins were used, to minimize non-specific labelling of the frozen sections of liver. With these techniques, the specific immune staining of RBP was observed within liver parenchymal cells. This staining appeared as both particulate and diffuse within the cytoplasm of the parenchymal cells, and was not concentrated within one region of the liver cell or lobule. Staining for RBP was not observed in nuclei or in cells other than parenchymal cells. Similar particulate and diffuse immune staining for RBP was observed in liver sections from both vitamin A-deficient and normal rats. More intense immune staining appeared to be present in the sections of vitamin A-deficient animals, in good correlation with the expected higher levels of RBP in deficient as compared to normal liver. When liver sections were exposed to an antiserum to rat albumin, instead of one to rat RBP, immune cytoplasmic staining was observed which was entirely of a diffuse nature, and did not appear particulate or granular. The findings suggest that RBP, unlike albumin, is localized in part within cytoplasmic vesicles or granules which are large enough to be detected with immunofluorescence, and which are present in livers of both normal and vitamin A-deficient animals. The nature of these putative RBP-containing particles remains to be explored.