<p>Orthoebolaviruses, like Ebola, Sudan, and Bundibugyo viruses, are responsible for causing sporadic and unpredictable outbreaks of severe disease throughout Sub-Saharan Africa. Accurate and reliable diagnostic tests are key to identifying cases of disease and restricting its spread, yet options are mostly limited to molecular assays, such as reverse transcriptase quantitative PCR, that require time, training, and complex equipment to run. To complement existing diagnostic modalities, and to address some of their shortcomings, we developed a prototype immunochromatographic lateral flow rapid diagnostic test (ILF-RDT) that detects the glycoproteins (GPs) from all known orthoebolaviruses, including Bundibugyo virus, which is currently causing a large outbreak in the Democratic Republic of the Congo and Uganda. This test depends on a pair of novel, broadly cross-reactive monoclonal antibodies capable of binding the viral GP—including the secreted GP—in a non-competitive manner. Here, we show that this ILF-RDT detects both recombinant GP and authentic virus in a variety of substrates, including samples spiked with human blood and blood samples taken from experimentally infected animals. Indeed, the ILF-RDT was able to detect Ebola virus in nonhuman primate samples at 4 days post-infection, before the animals began exhibiting signs of severe disease. Given the uniquely broad cross-reactivity of this diagnostic assay, its ability to detect low levels of virus early during infection, and its suitability for use at the point-of-care in resource-limited settings, this ILF-RDT could have a significant impact on the management and mitigation of future orthoebolavirus outbreaks.</p>