Activated monocytes forming intravascular clumps in the veins of most organs appeared in LEW rats after a 3-day intravenous treatment with recombinant rat gamma interferon. Phenotyping in situ and in cytospot preparations of perfusates revealed that the cells coexpressed the rat monocyte/macrophage antigen ED1 and class II MHC molecules. In addition, most cells reacted with a rat CD11b antibody and weakly expressed determinants detected by the W3/13 and Ox22 reagents. Minor fractions of the activated monocytes were positive for rat CD4 and the Ox2 and ED3 determinants. Cell proliferation was assessed by double staining for bromodeoxyuridine (BrdUrd) incorporation and phenotypic markers. Of the ED 1-positive class II-positive cells, 80% were labeled with BrdUrd after 3 days of combined infusion with gamma interferon. Pulse labeling for 30 minutes revealed 8% BrdUrd-positive intravascular ED 1-positive class II-positive monocytes in situ on day 3 of treatment, which contrasted with almost-absent labeling of this cell population in normal LEW rats. It is concluded that interferon not only promotes activation but also intravascular division of monocytes or their immediate precursors. Interestingly, cells of identical morphology and phenotype were observed in the vasculature of rats during lethal graft-versus-host reactions. Activated monocytes may thus contribute to the pathologic consequences of cytokine treatment and severe systemic immune reactions in vivo.