Lymphocyte traffic into lymph nodes and into mucosa-associated lymphoid tissues is regulated by specialized postcapillary high endothelial venules (HEVs). The authors have produced a rat monoclonal antibody, HECA-452, that detects a human endothelial cell differentiation antigen selectively expressed on high endothelium. In immunoperoxidase studies, HECA-452 intensely stains all HEVs within lymphoid organs. In normal nonlymphoid tissues the antibody stains no vascular endothelium. The antibody, in addition to reacting with high endothelium, cross-reacts with a set of monocytic cells. In pathologic states such as autoimmune thyroiditis and Crohn's disease, known for the development of dense, frequently organized, lymphocytic infiltrates, HECA-452 detects HEV-like vessels containing luminal and intramural lymphocytes, presumably in the process of extravasating. The antigen was not expressed at detectable levels by venules in less heavily infiltrated chronic inflammatory sites nor in acutely inflamed tissues. In lymphoid malignancies, the only vessels stained were morphologically characteristic HEVs in association with areas of residual normal lymphoid tissue or reactive lymphocytic infiltrates. The specificity of HECA-452 for high endothelial cells confirms the highly specialized nature of these vessels and will permit studies of the regulation of high endothelial cell differentiation in vivo and in vitro. The HECA-452 antigen is preserved in paraffin sections of sublimate formaldehyde- and also routinely formalin-fixed tissues. Thus, HECA-452 will be widely applicable for the immunohistologic detection of endothelium specialized for the support of highly increased lymphocyte extravasation in inflammatory sites.