The movement of proteins between the cytoplasm and nucleus mediated by the importin superfamily of proteins is an essential to many cellular processes including differentiation and development, and is critical to disease states such as viral disease and oncogenesis. We recently developed a high-throughput screen to identify specific and general inhibitors of protein nuclear import, from which Ivermectin was identified as a potential inhibitor of importin alpha/beta-mediated transport. Here we characterise in detail the nuclear transport inhibitory properties of Ivermectin, demonstrating that it is a broad-spectrum inhibitor of importin alpha/beta nuclear import with no effect on a range of other nuclear import pathways, including that mediated by importin beta1 alone. Importantly, we establish for the first time that Ivermectin has potent anti-viral activity towards both HIV-1 and Dengue virus, both of which are strongly reliant on importin alpha/beta nuclear import, with respect to the HIV-1 Integrase and NS5 polymerase proteins respectively. Ivermectin would appear to be an invaluable tool for the study of protein nuclear import, as well as the basis for future development of anti-viral agents.