In addition to preserving hemostasis, fibrinogen assembly on leukocytes mediates inflammatory responses and may aberrantly contribute to vascular injury. In this study, we used real-time intravital video microscopy in exposed rabbit mesentery to investigate the potential role of fibrinogen on leukocyte adherence mechanisms, in vivo. At physiologic concentrations of 0.15 to 0.5 mg/mL, human fibrinogen dose-dependently enhanced by threefold to fivefold the adhesion of chemoattractant-stimulated monocytic HL-60 cells to rabbit mesenteric endothelium, by acting as a bridging molecule between the two types. Fibrinogen-dependent intercellular bridging occurred in venules, but not in arterioles or capillaries (1), was optimal at reduced flow shear forces (range: 0.77 to 2.79 dyne/cm2) (2), and produced a firm attachment of monocytic cells to endothelium, rather than transient rolling (3). Consistent with this model, rabbit fibrinogen failed to support human leukocyte adhesion, while human fibrinogen enhanced monocytic cell attachment to rabbit endothelial cells in vitro, in a reaction indistinguishable from that observed with human endothelium. Antagonists of the recently described association of fibrinogen with intercellular adhesion molecule-1 (ICAM-1), including monoclonal antibodies (MoAbs) LB-2 or 2D5, or the fibrinogen gamma 3 peptide gamma Asn117-Ala133, blocked fibrinogen-dependent leukocyte-endothelium interaction in vitro or in vivo, respectively, while a control nonbinding antibody or the fibrinogen L10 peptide gamma Leu402-Val411 were ineffective. These data suggest that simultaneous assembly of fibrinogen on leukocytes and endothelial ICAM-1 provides a pathway of intercellular adhesion which may act in concert with beta 2 integrins to stabilize firm leukocyte attachment to endothelium, in vivo. Given the recognized role of fibrinogen as a major risk factor for atherosclerosis, this mechanism may directly contribute to thrombus formation and endothelial cell damage in vascular diseases.