Juvenile anguillid eels migrating into inland waters often face migration barriers. Upstream passage solutions normally consist of inclined ramps lined with a wetted climbing substrate. In this study, we compared the performance of three commonly used substrate types in a controlled experiment, using European eel as the test species. We also analyzed climbing behavior with videography and validated the experimental results under natural conditions at a hydropower plant. In addition, we investigated the effects of ramp placement. Studded substrate attracted more approaches and climbs and passed more eels at a higher climbing velocity than open weave and bristle substrates, results that were confirmed by the field validation. Moreover, ramps placed in the tailrace caught more eels in low than in high water velocities. To conserve anguillid eels, both safe routes for downstream-migrating adult silver eels and improved recruitment at the freshwater feeding life stage must be achieved. Optimizing ramp position and equipping upstream passage solutions with functioning climbing substrate are key factors to enhance the performance of eel ramps.