A new assessment tool, Ecology and Evolution--Measuring Achievement and Progression in Science or EcoEvo-MAPS, measures student thinking in ecology and evolution during an undergraduate course of study. EcoEvo-MAPS targets foundational concepts in ecology and evolution and uses a novel approach that asks students to evaluate a series of predictions, conclusions, or interpretations as likely or unlikely to be true given a specific scenario. We collected evidence of validity and reliability for EcoEvo-MAPS through an iterative process of faculty review, student interviews, and analyses of assessment data from more than 3000 students at 34 associate's-, bachelor's-, master's-, and doctoral-granting institutions. The 63 likely/unlikely statements range in difficulty and target student understanding of key concepts aligned with the "Vision and Change" report. This assessment provides departments with a tool to measure student thinking at different time points in the curriculum and provides data that can be used to inform curricular and instructional modifications.