The latest outbreak of Ebola virus disease (EVD) in West Africa has highlighted the urgent need for the development of rapid and reliable diagnostic assays. We used monoclonal antibodies specific to the ebolavirus nucleoprotein to develop an immunochromatography (IC) assay (QuickNavi-Ebola) for rapid diagnosis of EVD. The IC assay was first evaluated with tissue culture supernatants of infected Vero E6 cells and found to be capable of detecting 103–104 focus-forming units/mL of ebolaviruses. Using serum samples from experimentally infected nonhuman primates, we confirmed that the assay could detect the viral antigen shortly after disease onset. It was also noted that multiple species of ebolaviruses could be detected by the IC assay. Owing to the simplicity of the assay procedure and absence of requirements for special equipment and training, QuickNavi-Ebola is expected to be a useful tool for rapid diagnosis of EVD.