Disclosed in the present invention is a fishway effect evaluation method based on stable isotope method, belonging to the technical field of fishway effect evaluation. The fishway effect evaluation method according to the present invention comprises stable isotope ratio determination, trophic niche estimation and effectiveness evaluation on fishway construction. The evaluation method according to the present invention is used for researching one or more migratory fishes in the upstream and downstream of the water conservancy projects, determining and quantitatively explaining whether the influence of the water conservancy projects on one or more migratory fishes is eliminated or not from the perspective of the ecosystem through comparison and analysis of stable isotopes, and has the characteristics of high detection speed, stable result and the like.