The serious medical consequences and costly management of infections associated with vascular grafts have prompted an expanding interest in examining the preventive efficacy of antimicrobial-coated vascular grafts. The purpose of antimicrobial coating of vascular grafts is to reduce bacterial colonization of the device and, hopefully, the occurrence of clinical infection. In this study we demonstrated that expanded-polytetrafluoroethylene vascular grafts coated with minocycline and rifampin provide broad-spectrum antimicrobial activity in vitro, as reflected by zones of inhibition, against Staphylococcus epidermidis, S. aureus, Enterococcus faecium, and Pseudomonas aeruginosa. We also showed in a rabbit model that subcutaneously placed minocycline/rifampin-coated vascular grafts have lower rates of staphylococcal device colonization (1/24 = 4% vs. 8/30 = 27%, p = 0.033) and device-related infection (0/24 = 0% vs. 6/30 = 20%, p = 0.028) than uncoated grafts. These promising results encourage the clinical evaluation of vascular grafts coated with minocycline and rifampin.