AbstractGlobal warming is causing an unprecedented loss of species and habitats worldwide. This is particularly apparent for tropical coral reefs, with an increasing number of reefs experiencing mass bleaching and mortality on an annual basis. As such, there is a growing need for a standardized experimental approach to rapidly assess the thermal limits of corals and predict the survival of coral species across reefs and regions. Using a portable experimental system, the Coral Bleaching Automated Stress System (CBASS), we conducted standardized 18 h acute thermal stress assays to quantitively determine the upper thermal limits of four coral species across the length of the Red Sea coastline, from the Gulf of Aqaba (GoA) to Djibouti (~ 2100 km). We measured dark-acclimated photosynthetic efficiency (F_v/F_m), algal symbiont density, chlorophyll a, and visual bleaching intensity following heat stress. F_v/F_m was the most precise response variable assessed, advancing the F_v/F_m effective dose 50 (ED50, i.e., the temperature at which 50% of the initial F_v/F_m is measured) as an empirically derived proxy for thermal tolerance. ED50 thermal thresholds from the central/southern Red Sea and Djibouti populations were consistently higher for Acropora hemprichii, Pocillopora verrucosa, and Stylophora pistillata (0.1–1.8 °C above GoA corals, respectively), in line with prevailing warmer maximum monthly means (MMMs), though were lower than GoA corals relative to site MMMs (1.5–3.0 °C). P. verrucosa had the lowest thresholds overall. Despite coming from the hottest site, thresholds were lowest for Porites lobata in the southern Red Sea, suggesting long-term physiological damage or ongoing recovery from a severe, prior bleaching event. Altogether, the CBASS resolved historical, taxonomic, and possibly recent environmental drivers of variation in coral thermal thresholds, highlighting the potential for a standardized, short-term thermal assay as a universal approach for assessing ecological and evolutionary variation in the upper thermal limits of corals.