Endothelial-Monocyte-Activating Polypeptide II (EMAP II) is a proinflammatory cytokine and chemoattractant of macrophages. In order to investigate the role of EMAP II in autoimmune lesions of the rat nervous system, we have used a synthetic gene to express EMAP II in E. coli and have produced monoclonal antibodies against EMAP II. Monoclonal antibodies are suited to demonstrate EMAP II in ELISAs, Western blots, and paraffin-embedded tissue sections. EMAP II was localized to monocytes/macrophages with rather selective staining of a minor rat monocyte subpopulation of lymphoid tissues such as spleen, lymph nodes or follicles of the gut. In the normal brain, cells of the perivascular but not parenchymal microglia were stained. We then investigated expression of EMAP II during experimental autoimmune encephalomyelitis (EAE), neuritis (EAN), and uveitis (EAU). Within the local inflammatory lesions infiltrating macrophages are prominently stained. In the diseased brain, EMAP II-positive microglial cells are not only found in the direct vicinity of the inflammatory infiltrate, but widespread activation is seen in the parenchyma. This is the first demonstration that EMAP II is present in autoimmune lesions. Immunostaining of microglial cells is noteworthy, as these cells are strategically placed regulatory elements of CNS immunosurveillance. EMAP II might be a factor regulating monocyte chemoattraction, endothelial cell activation and a regulator of microglial cell reactivity in autoimmune inflammation of the central nervous system.